<?xml version="1.0" encoding="us-ascii"?>
<!-- DFIN ActiveDisclosure(SM) XBRL Linkbase Document - http://www.dfinsolutions.com/ -->
<!-- Creation Date      : 2022-08-08T16:42:56.7141743+00:00 -->
<!-- Version            : 5.0.1.747 -->
<!-- Package ID         : 4fc44d86180b4cdfb2d650485e0d2cd0 -->
<!-- Copyright (c) 2022 Donnelley Financial Solutions, Inc. All Rights Reserved. -->
<link:linkbase xmlns:link="http://www.xbrl.org/2003/linkbase" xmlns:xbrldt="http://xbrl.org/2005/xbrldt" xmlns:xlink="http://www.w3.org/1999/xlink" xmlns:xsi="http://www.w3.org/2001/XMLSchema-instance" xsi:schemaLocation="http://www.xbrl.org/2003/linkbase http://www.xbrl.org/2003/xbrl-linkbase-2003-12-31.xsd">
  <link:arcroleRef xlink:type="simple" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#all" arcroleURI="http://xbrl.org/int/dim/arcrole/all" />
  <link:arcroleRef xlink:type="simple" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#dimension-default" arcroleURI="http://xbrl.org/int/dim/arcrole/dimension-default" />
  <link:arcroleRef xlink:type="simple" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#dimension-domain" arcroleURI="http://xbrl.org/int/dim/arcrole/dimension-domain" />
  <link:arcroleRef xlink:type="simple" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#domain-member" arcroleURI="http://xbrl.org/int/dim/arcrole/domain-member" />
  <link:arcroleRef xlink:type="simple" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#hypercube-dimension" arcroleURI="http://xbrl.org/int/dim/arcrole/hypercube-dimension" />
  <link:roleRef xlink:type="simple" xlink:href="onto-20220702.xsd#TemplateLink" roleURI="http://www.ontoinnovation.com/20220702/role/TemplateLink" />
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/StatementCONDENSEDCONSOLIDATEDSTATEMENTSOFSTOCKHOLDERSEQUITYUnaudited" xlink:href="onto-20220702.xsd#Role_StatementCONDENSEDCONSOLIDATEDSTATEMENTSOFSTOCKHOLDERSEQUITYUnaudited" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/StatementCONDENSEDCONSOLIDATEDSTATEMENTSOFSTOCKHOLDERSEQUITYUnaudited">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_OtherComprehensiveIncomeUnrealizedHoldingGainLossOnSecuritiesArisingDuringPeriodNetOfTax" xlink:type="locator" xlink:label="us-gaap_OtherComprehensiveIncomeUnrealizedHoldingGainLossOnSecuritiesArisingDuringPeriodNetOfTax" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_OtherComprehensiveIncomeForeignCurrencyTransactionAndTranslationAdjustmentNetOfTaxPortionAttributableToParent" xlink:type="locator" xlink:label="us-gaap_OtherComprehensiveIncomeForeignCurrencyTransactionAndTranslationAdjustmentNetOfTaxPortionAttributableToParent" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_SharesPaidForTaxWithholdingForShareBasedCompensation" xlink:type="locator" xlink:label="us-gaap_SharesPaidForTaxWithholdingForShareBasedCompensation" />
    <link:loc xlink:href="onto-20220702.xsd#onto_AdjustmentsToAdditionalPaidInCapitalIncomeTaxDeficiencyFromShareBasedCompensations" xlink:type="locator" xlink:label="onto_AdjustmentsToAdditionalPaidInCapitalIncomeTaxDeficiencyFromShareBasedCompensations" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockIssuedDuringPeriodSharesShareBasedCompensation" xlink:type="locator" xlink:label="us-gaap_StockIssuedDuringPeriodSharesShareBasedCompensation" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockIssuedDuringPeriodValueShareBasedCompensation" xlink:type="locator" xlink:label="us-gaap_StockIssuedDuringPeriodValueShareBasedCompensation" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AdjustmentsToAdditionalPaidInCapitalSharebasedCompensationRequisiteServicePeriodRecognitionValue" xlink:type="locator" xlink:label="us-gaap_AdjustmentsToAdditionalPaidInCapitalSharebasedCompensationRequisiteServicePeriodRecognitionValue" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_NetIncomeLoss" xlink:type="locator" xlink:label="us-gaap_NetIncomeLoss" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CommonStockSharesIssued" xlink:type="locator" xlink:label="us-gaap_CommonStockSharesIssued" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockholdersEquity" xlink:type="locator" xlink:label="us-gaap_StockholdersEquity" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_RetainedEarningsMember" xlink:type="locator" xlink:label="us-gaap_RetainedEarningsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedOtherComprehensiveIncomeMember" xlink:type="locator" xlink:label="us-gaap_AccumulatedOtherComprehensiveIncomeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AdditionalPaidInCapitalMember" xlink:type="locator" xlink:label="us-gaap_AdditionalPaidInCapitalMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CommonStockMember" xlink:type="locator" xlink:label="us-gaap_CommonStockMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_EquityComponentDomain" xlink:type="locator" xlink:label="us-gaap_EquityComponentDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_EquityComponentDomain" xlink:type="locator" xlink:label="us-gaap_EquityComponentDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StatementEquityComponentsAxis" xlink:type="locator" xlink:label="us-gaap_StatementEquityComponentsAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StatementTable" xlink:type="locator" xlink:label="us-gaap_StatementTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StatementLineItems" xlink:type="locator" xlink:label="us-gaap_StatementLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_StatementTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_StatementTable" xlink:to="us-gaap_StatementEquityComponentsAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_StatementEquityComponentsAxis" xlink:to="us-gaap_EquityComponentDomain_2" order="10050.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_StatementEquityComponentsAxis" xlink:to="us-gaap_EquityComponentDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_CommonStockMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_AdditionalPaidInCapitalMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_AccumulatedOtherComprehensiveIncomeMember" order="10150.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_RetainedEarningsMember" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_StockholdersEquity" order="10340.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_CommonStockSharesIssued" order="10360.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_NetIncomeLoss" order="10480.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_AdjustmentsToAdditionalPaidInCapitalSharebasedCompensationRequisiteServicePeriodRecognitionValue" order="10620.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_StockIssuedDuringPeriodValueShareBasedCompensation" order="10760.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_StockIssuedDuringPeriodSharesShareBasedCompensation" order="10780.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="onto_AdjustmentsToAdditionalPaidInCapitalIncomeTaxDeficiencyFromShareBasedCompensations" order="10900.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_SharesPaidForTaxWithholdingForShareBasedCompensation" order="10920.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_OtherComprehensiveIncomeForeignCurrencyTransactionAndTranslationAdjustmentNetOfTaxPortionAttributableToParent" order="11040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_StatementLineItems" xlink:to="us-gaap_OtherComprehensiveIncomeUnrealizedHoldingGainLossOnSecuritiesArisingDuringPeriodNetOfTax" order="11180.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureBusinessCombinationTextualDetails" xlink:href="onto-20220702.xsd#Role_DisclosureBusinessCombinationTextualDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureBusinessCombinationTextualDetails">
    <link:loc xlink:href="onto-20220702.xsd#onto_BusinessCombinationPotentialEarnoutSubjectToPrincipalRemainingPeriod" xlink:type="locator" xlink:label="onto_BusinessCombinationPotentialEarnoutSubjectToPrincipalRemainingPeriod" />
    <link:loc xlink:href="onto-20220702.xsd#onto_BusinessCombinationPotentialEarnoutAccrued" xlink:type="locator" xlink:label="onto_BusinessCombinationPotentialEarnoutAccrued" />
    <link:loc xlink:href="onto-20220702.xsd#onto_BusinessCombinationPotentialEarnoutAdditionalPayment" xlink:type="locator" xlink:label="onto_BusinessCombinationPotentialEarnoutAdditionalPayment" />
    <link:loc xlink:href="onto-20220702.xsd#onto_BusinessAcquisitionAchievedEarnoutPayment" xlink:type="locator" xlink:label="onto_BusinessAcquisitionAchievedEarnoutPayment" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PaymentsToAcquireBusinessesGross" xlink:type="locator" xlink:label="us-gaap_PaymentsToAcquireBusinessesGross" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MinimumMember" xlink:type="locator" xlink:label="srt_MinimumMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MaximumMember" xlink:type="locator" xlink:label="srt_MaximumMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeMember" xlink:type="locator" xlink:label="srt_RangeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeMember" xlink:type="locator" xlink:label="srt_RangeMember_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeAxis" xlink:type="locator" xlink:label="srt_RangeAxis" />
    <link:loc xlink:href="onto-20220702.xsd#onto_InspectrologyLLCMember" xlink:type="locator" xlink:label="onto_InspectrologyLLCMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_BusinessAcquisitionAcquireeDomain" xlink:type="locator" xlink:label="us-gaap_BusinessAcquisitionAcquireeDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_BusinessAcquisitionAcquireeDomain" xlink:type="locator" xlink:label="us-gaap_BusinessAcquisitionAcquireeDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_BusinessAcquisitionAxis" xlink:type="locator" xlink:label="us-gaap_BusinessAcquisitionAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfBusinessAcquisitionsByAcquisitionTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfBusinessAcquisitionsByAcquisitionTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_BusinessAcquisitionLineItems" xlink:type="locator" xlink:label="us-gaap_BusinessAcquisitionLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_BusinessAcquisitionLineItems" xlink:to="us-gaap_ScheduleOfBusinessAcquisitionsByAcquisitionTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfBusinessAcquisitionsByAcquisitionTable" xlink:to="us-gaap_BusinessAcquisitionAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_BusinessAcquisitionAxis" xlink:to="us-gaap_BusinessAcquisitionAcquireeDomain_2" order="10050.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_BusinessAcquisitionAxis" xlink:to="us-gaap_BusinessAcquisitionAcquireeDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_BusinessAcquisitionAcquireeDomain" xlink:to="onto_InspectrologyLLCMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfBusinessAcquisitionsByAcquisitionTable" xlink:to="srt_RangeAxis" order="10200.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="srt_RangeAxis" xlink:to="srt_RangeMember_2" order="10210.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="srt_RangeAxis" xlink:to="srt_RangeMember" order="10230.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_RangeMember" xlink:to="srt_MaximumMember" order="10250.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_RangeMember" xlink:to="srt_MinimumMember" order="10270.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_BusinessAcquisitionLineItems" xlink:to="us-gaap_PaymentsToAcquireBusinessesGross" order="10320.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_BusinessAcquisitionLineItems" xlink:to="onto_BusinessAcquisitionAchievedEarnoutPayment" order="10340.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_BusinessAcquisitionLineItems" xlink:to="onto_BusinessCombinationPotentialEarnoutAdditionalPayment" order="10360.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_BusinessAcquisitionLineItems" xlink:to="onto_BusinessCombinationPotentialEarnoutAccrued" order="10380.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_BusinessAcquisitionLineItems" xlink:to="onto_BusinessCombinationPotentialEarnoutSubjectToPrincipalRemainingPeriod" order="10400.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureFairValueMeasurementsAssetsAndLiabilitiesCarriedAtFairValueMeasuredOnRecurringBasisDetails" xlink:href="onto-20220702.xsd#Role_DisclosureFairValueMeasurementsAssetsAndLiabilitiesCarriedAtFairValueMeasuredOnRecurringBasisDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureFairValueMeasurementsAssetsAndLiabilitiesCarriedAtFairValueMeasuredOnRecurringBasisDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_LiabilitiesFairValueDisclosure" xlink:type="locator" xlink:label="us-gaap_LiabilitiesFairValueDisclosure" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_BusinessCombinationContingentConsiderationLiability" xlink:type="locator" xlink:label="us-gaap_BusinessCombinationContingentConsiderationLiability" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ForeignCurrencyContractsLiabilityFairValueDisclosure" xlink:type="locator" xlink:label="us-gaap_ForeignCurrencyContractsLiabilityFairValueDisclosure" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AssetsFairValueDisclosure" xlink:type="locator" xlink:label="us-gaap_AssetsFairValueDisclosure" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AvailableForSaleSecuritiesDebtSecurities" xlink:type="locator" xlink:label="us-gaap_AvailableForSaleSecuritiesDebtSecurities" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CorporateDebtSecuritiesMember" xlink:type="locator" xlink:label="us-gaap_CorporateDebtSecuritiesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" xlink:type="locator" xlink:label="us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CertificatesOfDepositMember" xlink:type="locator" xlink:label="us-gaap_CertificatesOfDepositMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AssetBackedSecuritiesMember" xlink:type="locator" xlink:label="us-gaap_AssetBackedSecuritiesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_USStatesAndPoliticalSubdivisionsMember" xlink:type="locator" xlink:label="us-gaap_USStatesAndPoliticalSubdivisionsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:type="locator" xlink:label="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:type="locator" xlink:label="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueByAssetClassAxis" xlink:type="locator" xlink:label="us-gaap_FairValueByAssetClassAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueMeasurementsRecurringMember" xlink:type="locator" xlink:label="us-gaap_FairValueMeasurementsRecurringMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueMeasurementFrequencyDomain" xlink:type="locator" xlink:label="us-gaap_FairValueMeasurementFrequencyDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueMeasurementFrequencyDomain" xlink:type="locator" xlink:label="us-gaap_FairValueMeasurementFrequencyDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueByMeasurementFrequencyAxis" xlink:type="locator" xlink:label="us-gaap_FairValueByMeasurementFrequencyAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CarryingReportedAmountFairValueDisclosureMember" xlink:type="locator" xlink:label="us-gaap_CarryingReportedAmountFairValueDisclosureMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueDisclosureItemAmountsDomain" xlink:type="locator" xlink:label="us-gaap_FairValueDisclosureItemAmountsDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PortionAtFairValueFairValueDisclosureMember" xlink:type="locator" xlink:label="us-gaap_PortionAtFairValueFairValueDisclosureMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueByMeasurementBasisAxis" xlink:type="locator" xlink:label="us-gaap_FairValueByMeasurementBasisAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueInputsLevel3Member" xlink:type="locator" xlink:label="us-gaap_FairValueInputsLevel3Member" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueInputsLevel2Member" xlink:type="locator" xlink:label="us-gaap_FairValueInputsLevel2Member" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueInputsLevel1Member" xlink:type="locator" xlink:label="us-gaap_FairValueInputsLevel1Member" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueMeasurementsFairValueHierarchyDomain" xlink:type="locator" xlink:label="us-gaap_FairValueMeasurementsFairValueHierarchyDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueMeasurementsFairValueHierarchyDomain" xlink:type="locator" xlink:label="us-gaap_FairValueMeasurementsFairValueHierarchyDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueByFairValueHierarchyLevelAxis" xlink:type="locator" xlink:label="us-gaap_FairValueByFairValueHierarchyLevelAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" xlink:type="locator" xlink:label="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:type="locator" xlink:label="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:to="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" order="10080.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" xlink:to="us-gaap_FairValueByFairValueHierarchyLevelAxis" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_FairValueByFairValueHierarchyLevelAxis" xlink:to="us-gaap_FairValueMeasurementsFairValueHierarchyDomain_2" order="10120.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_FairValueByFairValueHierarchyLevelAxis" xlink:to="us-gaap_FairValueMeasurementsFairValueHierarchyDomain" order="10140.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueMeasurementsFairValueHierarchyDomain" xlink:to="us-gaap_FairValueInputsLevel1Member" order="10160.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueMeasurementsFairValueHierarchyDomain" xlink:to="us-gaap_FairValueInputsLevel2Member" order="10180.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueMeasurementsFairValueHierarchyDomain" xlink:to="us-gaap_FairValueInputsLevel3Member" order="10200.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" xlink:to="us-gaap_FairValueByMeasurementBasisAxis" order="10210.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_FairValueByMeasurementBasisAxis" xlink:to="us-gaap_PortionAtFairValueFairValueDisclosureMember" order="10220.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueDisclosureItemAmountsDomain" xlink:to="us-gaap_PortionAtFairValueFairValueDisclosureMember" order="10230.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_FairValueByMeasurementBasisAxis" xlink:to="us-gaap_FairValueDisclosureItemAmountsDomain" order="10260.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueDisclosureItemAmountsDomain" xlink:to="us-gaap_CarryingReportedAmountFairValueDisclosureMember" order="10280.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" xlink:to="us-gaap_FairValueByMeasurementFrequencyAxis" order="10290.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_FairValueByMeasurementFrequencyAxis" xlink:to="us-gaap_FairValueMeasurementFrequencyDomain_2" order="10300.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_FairValueByMeasurementFrequencyAxis" xlink:to="us-gaap_FairValueMeasurementFrequencyDomain" order="10320.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueMeasurementFrequencyDomain" xlink:to="us-gaap_FairValueMeasurementsRecurringMember" order="10340.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisTable" xlink:to="us-gaap_FairValueByAssetClassAxis" order="10410.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_FairValueByAssetClassAxis" xlink:to="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain_2" order="10420.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_FairValueByAssetClassAxis" xlink:to="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" order="10440.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:to="us-gaap_USStatesAndPoliticalSubdivisionsMember" order="10460.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:to="us-gaap_AssetBackedSecuritiesMember" order="10480.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:to="us-gaap_CertificatesOfDepositMember" order="10500.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:to="us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" order="10520.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsMeasuredOnRecurringBasisUnobservableInputReconciliationByAssetClassDomain" xlink:to="us-gaap_CorporateDebtSecuritiesMember" order="10540.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:to="us-gaap_AvailableForSaleSecuritiesDebtSecurities" order="10670.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:to="us-gaap_AssetsFairValueDisclosure" order="11170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:to="us-gaap_ForeignCurrencyContractsLiabilityFairValueDisclosure" order="11270.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:to="us-gaap_BusinessCombinationContingentConsiderationLiability" order="11370.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FairValueAssetsAndLiabilitiesMeasuredOnRecurringAndNonrecurringBasisLineItems" xlink:to="us-gaap_LiabilitiesFairValueDisclosure" order="11470.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureMarketableSecuritiesScheduleOfMarketableSecuritiesByCategoryDetails" xlink:href="onto-20220702.xsd#Role_DisclosureMarketableSecuritiesScheduleOfMarketableSecuritiesByCategoryDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureMarketableSecuritiesScheduleOfMarketableSecuritiesByCategoryDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AvailableForSaleSecuritiesDebtSecurities" xlink:type="locator" xlink:label="us-gaap_AvailableForSaleSecuritiesDebtSecurities" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AvailableForSaleDebtSecuritiesAccumulatedGrossUnrealizedLossBeforeTax" xlink:type="locator" xlink:label="us-gaap_AvailableForSaleDebtSecuritiesAccumulatedGrossUnrealizedLossBeforeTax" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AvailableForSaleDebtSecuritiesAccumulatedGrossUnrealizedGainBeforeTax" xlink:type="locator" xlink:label="us-gaap_AvailableForSaleDebtSecuritiesAccumulatedGrossUnrealizedGainBeforeTax" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AvailableForSaleDebtSecuritiesAmortizedCostBasis" xlink:type="locator" xlink:label="us-gaap_AvailableForSaleDebtSecuritiesAmortizedCostBasis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CorporateDebtSecuritiesMember" xlink:type="locator" xlink:label="us-gaap_CorporateDebtSecuritiesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" xlink:type="locator" xlink:label="us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CertificatesOfDepositMember" xlink:type="locator" xlink:label="us-gaap_CertificatesOfDepositMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AssetBackedSecuritiesMember" xlink:type="locator" xlink:label="us-gaap_AssetBackedSecuritiesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_USTreasuryAndGovernmentMember" xlink:type="locator" xlink:label="us-gaap_USTreasuryAndGovernmentMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_InvestmentTypeCategorizationMember" xlink:type="locator" xlink:label="us-gaap_InvestmentTypeCategorizationMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_InvestmentTypeCategorizationMember" xlink:type="locator" xlink:label="us-gaap_InvestmentTypeCategorizationMember_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_InvestmentTypeAxis" xlink:type="locator" xlink:label="us-gaap_InvestmentTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:type="locator" xlink:label="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" order="10040.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" xlink:to="us-gaap_InvestmentTypeAxis" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_InvestmentTypeAxis" xlink:to="us-gaap_InvestmentTypeCategorizationMember_2" order="10080.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_InvestmentTypeAxis" xlink:to="us-gaap_InvestmentTypeCategorizationMember" order="10100.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_USTreasuryAndGovernmentMember" order="10120.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_AssetBackedSecuritiesMember" order="10140.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_CertificatesOfDepositMember" order="10160.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" order="10180.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_CorporateDebtSecuritiesMember" order="10200.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_AvailableForSaleDebtSecuritiesAmortizedCostBasis" order="10330.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_AvailableForSaleDebtSecuritiesAccumulatedGrossUnrealizedGainBeforeTax" order="10570.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_AvailableForSaleDebtSecuritiesAccumulatedGrossUnrealizedLossBeforeTax" order="10810.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_AvailableForSaleSecuritiesDebtSecurities" order="11050.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureMarketableSecuritiesSummaryOfEstimatedFairValueAndGrossUnrealizedHoldingLossesOfMarketableSecuritiesInUnrealizedLossPositionDetails" xlink:href="onto-20220702.xsd#Role_DisclosureMarketableSecuritiesSummaryOfEstimatedFairValueAndGrossUnrealizedHoldingLossesOfMarketableSecuritiesInUnrealizedLossPositionDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureMarketableSecuritiesSummaryOfEstimatedFairValueAndGrossUnrealizedHoldingLossesOfMarketableSecuritiesInUnrealizedLossPositionDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPosition12MonthsOrLongerAccumulatedLoss" xlink:type="locator" xlink:label="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPosition12MonthsOrLongerAccumulatedLoss" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPosition12MonthsOrLonger" xlink:type="locator" xlink:label="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPosition12MonthsOrLonger" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPositionLessThan12MonthsAccumulatedLoss" xlink:type="locator" xlink:label="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPositionLessThan12MonthsAccumulatedLoss" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPositionLessThan12Months" xlink:type="locator" xlink:label="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPositionLessThan12Months" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CorporateDebtSecuritiesMember" xlink:type="locator" xlink:label="us-gaap_CorporateDebtSecuritiesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" xlink:type="locator" xlink:label="us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CertificatesOfDepositMember" xlink:type="locator" xlink:label="us-gaap_CertificatesOfDepositMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AssetBackedSecuritiesMember" xlink:type="locator" xlink:label="us-gaap_AssetBackedSecuritiesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_USTreasuryAndGovernmentMember" xlink:type="locator" xlink:label="us-gaap_USTreasuryAndGovernmentMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_InvestmentTypeCategorizationMember" xlink:type="locator" xlink:label="us-gaap_InvestmentTypeCategorizationMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_InvestmentTypeAxis" xlink:type="locator" xlink:label="us-gaap_InvestmentTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:type="locator" xlink:label="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesTable" xlink:to="us-gaap_InvestmentTypeAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_InvestmentTypeAxis" xlink:to="us-gaap_InvestmentTypeCategorizationMember" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_USTreasuryAndGovernmentMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_AssetBackedSecuritiesMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_CertificatesOfDepositMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_CommercialPaperNotIncludedWithCashAndCashEquivalentsMember" order="10150.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_InvestmentTypeCategorizationMember" xlink:to="us-gaap_CorporateDebtSecuritiesMember" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPositionLessThan12Months" order="10280.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPositionLessThan12MonthsAccumulatedLoss" order="10500.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPosition12MonthsOrLonger" order="10720.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ScheduleOfAvailableForSaleSecuritiesLineItems" xlink:to="us-gaap_DebtSecuritiesAvailableForSaleContinuousUnrealizedLossPosition12MonthsOrLongerAccumulatedLoss" order="10940.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosurePurchasedIntangibleAssetsScheduleOfPurchasedIntangibleAssetsDetails" xlink:href="onto-20220702.xsd#Role_DisclosurePurchasedIntangibleAssetsScheduleOfPurchasedIntangibleAssetsDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosurePurchasedIntangibleAssetsScheduleOfPurchasedIntangibleAssetsDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_IntangibleAssetsNetExcludingGoodwill" xlink:type="locator" xlink:label="us-gaap_IntangibleAssetsNetExcludingGoodwill" />
    <link:loc xlink:href="onto-20220702.xsd#onto_IntangibleAssetsAccumulatedAmortizationExcludingGoodwill" xlink:type="locator" xlink:label="onto_IntangibleAssetsAccumulatedAmortizationExcludingGoodwill" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_IntangibleAssetsGrossExcludingGoodwill" xlink:type="locator" xlink:label="us-gaap_IntangibleAssetsGrossExcludingGoodwill" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsNet" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsNet" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsAccumulatedAmortization" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsAccumulatedAmortization" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsGross" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsGross" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_TrademarksAndTradeNamesMember" xlink:type="locator" xlink:label="us-gaap_TrademarksAndTradeNamesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CustomerRelationshipsMember" xlink:type="locator" xlink:label="us-gaap_CustomerRelationshipsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DevelopedTechnologyRightsMember" xlink:type="locator" xlink:label="us-gaap_DevelopedTechnologyRightsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsByMajorClassAxis" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsByMajorClassAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfFiniteLivedIntangibleAssetsTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfFiniteLivedIntangibleAssetsTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:type="locator" xlink:label="us-gaap_FiniteLivedIntangibleAssetsLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="us-gaap_ScheduleOfFiniteLivedIntangibleAssetsTable" order="10050.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfFiniteLivedIntangibleAssetsTable" xlink:to="us-gaap_FiniteLivedIntangibleAssetsByMajorClassAxis" order="10080.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_FiniteLivedIntangibleAssetsByMajorClassAxis" xlink:to="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain_2" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_FiniteLivedIntangibleAssetsByMajorClassAxis" xlink:to="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" xlink:to="us-gaap_DevelopedTechnologyRightsMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" xlink:to="us-gaap_CustomerRelationshipsMember" order="10150.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsMajorClassNameDomain" xlink:to="us-gaap_TrademarksAndTradeNamesMember" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="us-gaap_FiniteLivedIntangibleAssetsGross" order="10420.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="us-gaap_FiniteLivedIntangibleAssetsAccumulatedAmortization" order="10580.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="us-gaap_FiniteLivedIntangibleAssetsNet" order="10740.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="us-gaap_IntangibleAssetsGrossExcludingGoodwill" order="10930.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="onto_IntangibleAssetsAccumulatedAmortizationExcludingGoodwill" order="11090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_FiniteLivedIntangibleAssetsLineItems" xlink:to="us-gaap_IntangibleAssetsNetExcludingGoodwill" order="11250.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureBalanceSheetDetailsScheduleOfPropertyPlantAndEquipmentNetDetails" xlink:href="onto-20220702.xsd#Role_DisclosureBalanceSheetDetailsScheduleOfPropertyPlantAndEquipmentNetDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureBalanceSheetDetailsScheduleOfPropertyPlantAndEquipmentNetDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PropertyPlantAndEquipmentNet" xlink:type="locator" xlink:label="us-gaap_PropertyPlantAndEquipmentNet" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedDepreciationDepletionAndAmortizationPropertyPlantAndEquipment" xlink:type="locator" xlink:label="us-gaap_AccumulatedDepreciationDepletionAndAmortizationPropertyPlantAndEquipment" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PropertyPlantAndEquipmentGross" xlink:type="locator" xlink:label="us-gaap_PropertyPlantAndEquipmentGross" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_LeaseholdImprovementsMember" xlink:type="locator" xlink:label="us-gaap_LeaseholdImprovementsMember" />
    <link:loc xlink:href="onto-20220702.xsd#onto_ComputerEquipmentAndSoftwareMember" xlink:type="locator" xlink:label="onto_ComputerEquipmentAndSoftwareMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_FurnitureAndFixturesMember" xlink:type="locator" xlink:label="us-gaap_FurnitureAndFixturesMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_MachineryAndEquipmentMember" xlink:type="locator" xlink:label="us-gaap_MachineryAndEquipmentMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_LandAndBuildingMember" xlink:type="locator" xlink:label="us-gaap_LandAndBuildingMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:type="locator" xlink:label="us-gaap_PropertyPlantAndEquipmentTypeDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:type="locator" xlink:label="us-gaap_PropertyPlantAndEquipmentTypeDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PropertyPlantAndEquipmentByTypeAxis" xlink:type="locator" xlink:label="us-gaap_PropertyPlantAndEquipmentByTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfPropertyPlantAndEquipmentTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfPropertyPlantAndEquipmentTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_PropertyPlantAndEquipmentLineItems" xlink:type="locator" xlink:label="us-gaap_PropertyPlantAndEquipmentLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_PropertyPlantAndEquipmentLineItems" xlink:to="us-gaap_ScheduleOfPropertyPlantAndEquipmentTable" order="10030.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfPropertyPlantAndEquipmentTable" xlink:to="us-gaap_PropertyPlantAndEquipmentByTypeAxis" order="10060.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_PropertyPlantAndEquipmentByTypeAxis" xlink:to="us-gaap_PropertyPlantAndEquipmentTypeDomain_2" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_PropertyPlantAndEquipmentByTypeAxis" xlink:to="us-gaap_PropertyPlantAndEquipmentTypeDomain" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:to="us-gaap_LandAndBuildingMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:to="us-gaap_MachineryAndEquipmentMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:to="us-gaap_FurnitureAndFixturesMember" order="10150.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:to="onto_ComputerEquipmentAndSoftwareMember" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:to="us-gaap_LeaseholdImprovementsMember" order="10190.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentLineItems" xlink:to="us-gaap_PropertyPlantAndEquipmentGross" order="10220.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentLineItems" xlink:to="us-gaap_AccumulatedDepreciationDepletionAndAmortizationPropertyPlantAndEquipment" order="10460.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_PropertyPlantAndEquipmentLineItems" xlink:to="us-gaap_PropertyPlantAndEquipmentNet" order="10500.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://fasb.org/us-gaap/role/eedm/ExtensibleEnumerationLists" xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-roles-2022.xsd#eedm" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://fasb.org/us-gaap/role/eedm/ExtensibleEnumerationLists">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_OtherLiabilitiesNoncurrent" xlink:type="locator" xlink:label="us-gaap_OtherLiabilitiesNoncurrent" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_LiabilitiesNoncurrentAbstract" xlink:type="locator" xlink:label="us-gaap_LiabilitiesNoncurrentAbstract" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_LiabilitiesNoncurrentAbstract" xlink:to="us-gaap_OtherLiabilitiesNoncurrent" order="10150.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureCommitmentsAndContingenciesTextualDetails" xlink:href="onto-20220702.xsd#Role_DisclosureCommitmentsAndContingenciesTextualDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureCommitmentsAndContingenciesTextualDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_LineOfCreditFacilityInterestRateAtPeriodEnd" xlink:type="locator" xlink:label="us-gaap_LineOfCreditFacilityInterestRateAtPeriodEnd" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_LineOfCreditFacilityRemainingBorrowingCapacity" xlink:type="locator" xlink:label="us-gaap_LineOfCreditFacilityRemainingBorrowingCapacity" />
    <link:loc xlink:href="onto-20220702.xsd#onto_MaximumBorrowingCapacityPercentageOfEligibleSecuritiesHeldAtLineOfCreditAccessed" xlink:type="locator" xlink:label="onto_MaximumBorrowingCapacityPercentageOfEligibleSecuritiesHeldAtLineOfCreditAccessed" />
    <link:loc xlink:href="onto-20220702.xsd#onto_ProductWarrantyAccrualWarrantyPeriod" xlink:type="locator" xlink:label="onto_ProductWarrantyAccrualWarrantyPeriod" />
    <link:loc xlink:href="onto-20220702.xsd#onto_IntellectualPropertyIndemnificationObligations" xlink:type="locator" xlink:label="onto_IntellectualPropertyIndemnificationObligations" />
    <link:loc xlink:href="onto-20220702.xsd#onto_DueFromUnrelatedThirdParties" xlink:type="locator" xlink:label="onto_DueFromUnrelatedThirdParties" />
    <link:loc xlink:href="onto-20220702.xsd#onto_ProceedsFromAccountsReceivable" xlink:type="locator" xlink:label="onto_ProceedsFromAccountsReceivable" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MaximumMember" xlink:type="locator" xlink:label="srt_MaximumMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MinimumMember" xlink:type="locator" xlink:label="srt_MinimumMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeMember" xlink:type="locator" xlink:label="srt_RangeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeAxis" xlink:type="locator" xlink:label="srt_RangeAxis" />
    <link:loc xlink:href="onto-20220702.xsd#onto_CommitmentsAndContingenciesTable" xlink:type="locator" xlink:label="onto_CommitmentsAndContingenciesTable" />
    <link:loc xlink:href="onto-20220702.xsd#onto_CommitmentsAndContingenciesLineItems" xlink:type="locator" xlink:label="onto_CommitmentsAndContingenciesLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="onto_CommitmentsAndContingenciesTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="onto_CommitmentsAndContingenciesTable" xlink:to="srt_RangeAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="srt_RangeAxis" xlink:to="srt_RangeMember" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_RangeMember" xlink:to="srt_MinimumMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_RangeMember" xlink:to="srt_MaximumMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="onto_ProceedsFromAccountsReceivable" order="10140.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="onto_DueFromUnrelatedThirdParties" order="10160.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="onto_IntellectualPropertyIndemnificationObligations" order="10210.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="onto_ProductWarrantyAccrualWarrantyPeriod" order="10260.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="onto_MaximumBorrowingCapacityPercentageOfEligibleSecuritiesHeldAtLineOfCreditAccessed" order="10410.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="us-gaap_LineOfCreditFacilityRemainingBorrowingCapacity" order="10430.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_CommitmentsAndContingenciesLineItems" xlink:to="us-gaap_LineOfCreditFacilityInterestRateAtPeriodEnd" order="10450.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureRevenueScheduleOfDisaggregationOfRevenueByTimingOfRevenueDetails" xlink:href="onto-20220702.xsd#Role_DisclosureRevenueScheduleOfDisaggregationOfRevenueByTimingOfRevenueDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureRevenueScheduleOfDisaggregationOfRevenueByTimingOfRevenueDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" xlink:type="locator" xlink:label="us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_TransferredOverTimeMember" xlink:type="locator" xlink:label="us-gaap_TransferredOverTimeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_TransferredAtPointInTimeMember" xlink:type="locator" xlink:label="us-gaap_TransferredAtPointInTimeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_TimingOfTransferOfGoodOrServiceDomain" xlink:type="locator" xlink:label="us-gaap_TimingOfTransferOfGoodOrServiceDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_TimingOfTransferOfGoodOrServiceDomain" xlink:type="locator" xlink:label="us-gaap_TimingOfTransferOfGoodOrServiceDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_TimingOfTransferOfGoodOrServiceAxis" xlink:type="locator" xlink:label="us-gaap_TimingOfTransferOfGoodOrServiceAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DisaggregationOfRevenueTable" xlink:type="locator" xlink:label="us-gaap_DisaggregationOfRevenueTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_DisaggregationOfRevenueLineItems" xlink:type="locator" xlink:label="us-gaap_DisaggregationOfRevenueLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_DisaggregationOfRevenueLineItems" xlink:to="us-gaap_DisaggregationOfRevenueTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_DisaggregationOfRevenueTable" xlink:to="us-gaap_TimingOfTransferOfGoodOrServiceAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_TimingOfTransferOfGoodOrServiceAxis" xlink:to="us-gaap_TimingOfTransferOfGoodOrServiceDomain_2" order="10050.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_TimingOfTransferOfGoodOrServiceAxis" xlink:to="us-gaap_TimingOfTransferOfGoodOrServiceDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_TimingOfTransferOfGoodOrServiceDomain" xlink:to="us-gaap_TransferredAtPointInTimeMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_TimingOfTransferOfGoodOrServiceDomain" xlink:to="us-gaap_TransferredOverTimeMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_DisaggregationOfRevenueLineItems" xlink:to="us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" order="10140.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureShareBasedCompensationSummaryOfRestrictedStockUnitActivityDetails" xlink:href="onto-20220702.xsd#Role_DisclosureShareBasedCompensationSummaryOfRestrictedStockUnitActivityDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureShareBasedCompensationSummaryOfRestrictedStockUnitActivityDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeituresWeightedAverageGrantDateFairValue" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeituresWeightedAverageGrantDateFairValue" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriodWeightedAverageGrantDateFairValue" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriodWeightedAverageGrantDateFairValue" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriodWeightedAverageGrantDateFairValue" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriodWeightedAverageGrantDateFairValue" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeitedInPeriod" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeitedInPeriod" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriod" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriod" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriod" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriod" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_RestrictedStockUnitsRSUMember" xlink:type="locator" xlink:label="us-gaap_RestrictedStockUnitsRSUMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AwardTypeAxis" xlink:type="locator" xlink:label="us-gaap_AwardTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:to="us-gaap_AwardTypeAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_AwardTypeAxis" xlink:to="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2" order="10050.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_AwardTypeAxis" xlink:to="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:to="us-gaap_RestrictedStockUnitsRSUMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" order="10180.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" order="10300.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriod" order="10360.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriod" order="10420.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeitedInPeriod" order="10480.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" order="10630.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" order="10750.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriodWeightedAverageGrantDateFairValue" order="10810.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriodWeightedAverageGrantDateFairValue" order="10870.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeituresWeightedAverageGrantDateFairValue" order="10930.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureShareBasedCompensationTextualDetails" xlink:href="onto-20220702.xsd#Role_DisclosureShareBasedCompensationTextualDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureShareBasedCompensationTextualDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedPeriodForRecognition1" xlink:type="locator" xlink:label="us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedPeriodForRecognition1" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognized" xlink:type="locator" xlink:label="us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognized" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" />
    <link:loc xlink:href="onto-20220702.xsd#onto_PerformanceRestrictedStockUnitsMember" xlink:type="locator" xlink:label="onto_PerformanceRestrictedStockUnitsMember" />
    <link:loc xlink:href="onto-20220702.xsd#onto_RestrictedStockUnitsServiceBasedRSUsMember" xlink:type="locator" xlink:label="onto_RestrictedStockUnitsServiceBasedRSUsMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_RestrictedStockUnitsRSUMember" xlink:type="locator" xlink:label="us-gaap_RestrictedStockUnitsRSUMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AwardTypeAxis" xlink:type="locator" xlink:label="us-gaap_AwardTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:type="locator" xlink:label="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:to="us-gaap_AwardTypeAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_AwardTypeAxis" xlink:to="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:to="us-gaap_RestrictedStockUnitsRSUMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:to="onto_RestrictedStockUnitsServiceBasedRSUsMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:to="onto_PerformanceRestrictedStockUnitsMember" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" order="10220.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" order="10340.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognized" order="10380.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:to="us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedPeriodForRecognition1" order="10420.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureAccumulatedOtherComprehensiveIncomeLossComponentsOfAccumulatedOtherComprehensiveLossNetOfTaxDetails" xlink:href="onto-20220702.xsd#Role_DisclosureAccumulatedOtherComprehensiveIncomeLossComponentsOfAccumulatedOtherComprehensiveLossNetOfTaxDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureAccumulatedOtherComprehensiveIncomeLossComponentsOfAccumulatedOtherComprehensiveLossNetOfTaxDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ReclassificationFromAociCurrentPeriodNetOfTaxAttributableToParent" xlink:type="locator" xlink:label="us-gaap_ReclassificationFromAociCurrentPeriodNetOfTaxAttributableToParent" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_OciBeforeReclassificationsNetOfTaxAttributableToParent" xlink:type="locator" xlink:label="us-gaap_OciBeforeReclassificationsNetOfTaxAttributableToParent" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockholdersEquity" xlink:type="locator" xlink:label="us-gaap_StockholdersEquity" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedOtherComprehensiveIncomeMember" xlink:type="locator" xlink:label="us-gaap_AccumulatedOtherComprehensiveIncomeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedNetUnrealizedInvestmentGainLossMember" xlink:type="locator" xlink:label="us-gaap_AccumulatedNetUnrealizedInvestmentGainLossMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedTranslationAdjustmentMember" xlink:type="locator" xlink:label="us-gaap_AccumulatedTranslationAdjustmentMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_EquityComponentDomain" xlink:type="locator" xlink:label="us-gaap_EquityComponentDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StatementEquityComponentsAxis" xlink:type="locator" xlink:label="us-gaap_StatementEquityComponentsAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedOtherComprehensiveIncomeLossTable" xlink:type="locator" xlink:label="us-gaap_AccumulatedOtherComprehensiveIncomeLossTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_AccumulatedOtherComprehensiveIncomeLossLineItems" xlink:type="locator" xlink:label="us-gaap_AccumulatedOtherComprehensiveIncomeLossLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_AccumulatedOtherComprehensiveIncomeLossLineItems" xlink:to="us-gaap_AccumulatedOtherComprehensiveIncomeLossTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_AccumulatedOtherComprehensiveIncomeLossTable" xlink:to="us-gaap_StatementEquityComponentsAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_StatementEquityComponentsAxis" xlink:to="us-gaap_EquityComponentDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_AccumulatedTranslationAdjustmentMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_AccumulatedNetUnrealizedInvestmentGainLossMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_EquityComponentDomain" xlink:to="us-gaap_AccumulatedOtherComprehensiveIncomeMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_AccumulatedOtherComprehensiveIncomeLossLineItems" xlink:to="us-gaap_StockholdersEquity" order="10160.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_AccumulatedOtherComprehensiveIncomeLossLineItems" xlink:to="us-gaap_OciBeforeReclassificationsNetOfTaxAttributableToParent" order="10240.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_AccumulatedOtherComprehensiveIncomeLossLineItems" xlink:to="us-gaap_ReclassificationFromAociCurrentPeriodNetOfTaxAttributableToParent" order="10320.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueFromExternalCustomersByProductsAndServicesDetails" xlink:href="onto-20220702.xsd#Role_DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueFromExternalCustomersByProductsAndServicesDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueFromExternalCustomersByProductsAndServicesDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskPercentage1" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskPercentage1" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" xlink:type="locator" xlink:label="us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ProductConcentrationRiskMember" xlink:type="locator" xlink:label="us-gaap_ProductConcentrationRiskMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskTypeDomain" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskTypeDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskTypeDomain" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskTypeDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskByTypeAxis" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskByTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_SalesRevenueNetMember" xlink:type="locator" xlink:label="us-gaap_SalesRevenueNetMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskBenchmarkDomain" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskBenchmarkDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskBenchmarkDomain" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskBenchmarkDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskByBenchmarkAxis" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskByBenchmarkAxis" />
    <link:loc xlink:href="onto-20220702.xsd#onto_ServiceRevenueMember" xlink:type="locator" xlink:label="onto_ServiceRevenueMember" />
    <link:loc xlink:href="onto-20220702.xsd#onto_PartsRevenueMember" xlink:type="locator" xlink:label="onto_PartsRevenueMember" />
    <link:loc xlink:href="onto-20220702.xsd#onto_SystemsAndSoftwareRevenueMember" xlink:type="locator" xlink:label="onto_SystemsAndSoftwareRevenueMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_ProductsAndServicesDomain" xlink:type="locator" xlink:label="srt_ProductsAndServicesDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_ProductsAndServicesDomain" xlink:type="locator" xlink:label="srt_ProductsAndServicesDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_ProductOrServiceAxis" xlink:type="locator" xlink:label="srt_ProductOrServiceAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_SegmentReportingInformationLineItems" xlink:type="locator" xlink:label="us-gaap_SegmentReportingInformationLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_SegmentReportingInformationLineItems" xlink:to="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" xlink:to="srt_ProductOrServiceAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="srt_ProductOrServiceAxis" xlink:to="srt_ProductsAndServicesDomain_2" order="10050.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="srt_ProductOrServiceAxis" xlink:to="srt_ProductsAndServicesDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_ProductsAndServicesDomain" xlink:to="onto_SystemsAndSoftwareRevenueMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_ProductsAndServicesDomain" xlink:to="onto_PartsRevenueMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_ProductsAndServicesDomain" xlink:to="onto_ServiceRevenueMember" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" xlink:to="us-gaap_ConcentrationRiskByBenchmarkAxis" order="10140.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_ConcentrationRiskByBenchmarkAxis" xlink:to="us-gaap_ConcentrationRiskBenchmarkDomain_2" order="10150.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_ConcentrationRiskByBenchmarkAxis" xlink:to="us-gaap_ConcentrationRiskBenchmarkDomain" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ConcentrationRiskBenchmarkDomain" xlink:to="us-gaap_SalesRevenueNetMember" order="10190.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" xlink:to="us-gaap_ConcentrationRiskByTypeAxis" order="10260.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="us-gaap_ConcentrationRiskByTypeAxis" xlink:to="us-gaap_ConcentrationRiskTypeDomain_2" order="10270.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_ConcentrationRiskByTypeAxis" xlink:to="us-gaap_ConcentrationRiskTypeDomain" order="10290.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ConcentrationRiskTypeDomain" xlink:to="us-gaap_ProductConcentrationRiskMember" order="10310.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_SegmentReportingInformationLineItems" xlink:to="us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" order="10500.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_SegmentReportingInformationLineItems" xlink:to="us-gaap_ConcentrationRiskPercentage1" order="11090.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueFromExternalCustomersAttributedToForeignCountriesByGeographicAreaDetails" xlink:href="onto-20220702.xsd#Role_DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueFromExternalCustomersAttributedToForeignCountriesByGeographicAreaDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueFromExternalCustomersAttributedToForeignCountriesByGeographicAreaDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" xlink:type="locator" xlink:label="us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" />
    <link:loc xlink:href="https://xbrl.sec.gov/country/2022/country-2022.xsd#country_JP" xlink:type="locator" xlink:label="country_JP" />
    <link:loc xlink:href="onto-20220702.xsd#onto_SoutheastAsiaMember" xlink:type="locator" xlink:label="onto_SoutheastAsiaMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_EuropeMember" xlink:type="locator" xlink:label="srt_EuropeMember" />
    <link:loc xlink:href="https://xbrl.sec.gov/country/2022/country-2022.xsd#country_US" xlink:type="locator" xlink:label="country_US" />
    <link:loc xlink:href="https://xbrl.sec.gov/country/2022/country-2022.xsd#country_TW" xlink:type="locator" xlink:label="country_TW" />
    <link:loc xlink:href="https://xbrl.sec.gov/country/2022/country-2022.xsd#country_KR" xlink:type="locator" xlink:label="country_KR" />
    <link:loc xlink:href="https://xbrl.sec.gov/country/2022/country-2022.xsd#country_CN" xlink:type="locator" xlink:label="country_CN" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_SegmentGeographicalDomain" xlink:type="locator" xlink:label="srt_SegmentGeographicalDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_SegmentGeographicalDomain" xlink:type="locator" xlink:label="srt_SegmentGeographicalDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_StatementGeographicalAxis" xlink:type="locator" xlink:label="srt_StatementGeographicalAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" xlink:type="locator" xlink:label="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_SegmentReportingInformationLineItems" xlink:type="locator" xlink:label="us-gaap_SegmentReportingInformationLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_SegmentReportingInformationLineItems" xlink:to="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ScheduleOfSegmentReportingInformationBySegmentTable" xlink:to="srt_StatementGeographicalAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="srt_StatementGeographicalAxis" xlink:to="srt_SegmentGeographicalDomain_2" order="10050.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="srt_StatementGeographicalAxis" xlink:to="srt_SegmentGeographicalDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="country_CN" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="country_KR" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="country_TW" order="10130.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="country_US" order="10150.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="srt_EuropeMember" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="onto_SoutheastAsiaMember" order="10190.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_SegmentGeographicalDomain" xlink:to="country_JP" order="10210.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_SegmentReportingInformationLineItems" xlink:to="us-gaap_RevenueFromContractWithCustomerExcludingAssessedTax" order="10240.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueByMajorCustomerByReportingSegmentsDetails" xlink:href="onto-20220702.xsd#Role_DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueByMajorCustomerByReportingSegmentsDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureSegmentReportingAndGeographicInformationScheduleOfRevenueByMajorCustomerByReportingSegmentsDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskPercentage1" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskPercentage1" />
    <link:loc xlink:href="onto-20220702.xsd#onto_SamsungSemiconductorMember" xlink:type="locator" xlink:label="onto_SamsungSemiconductorMember" />
    <link:loc xlink:href="onto-20220702.xsd#onto_TaiwanSemiconductorManufacturingCompanyLimitedMember" xlink:type="locator" xlink:label="onto_TaiwanSemiconductorManufacturingCompanyLimitedMember" />
    <link:loc xlink:href="onto-20220702.xsd#onto_SKHynixIncMember" xlink:type="locator" xlink:label="onto_SKHynixIncMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_NameOfMajorCustomerDomain" xlink:type="locator" xlink:label="srt_NameOfMajorCustomerDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_NameOfMajorCustomerDomain" xlink:type="locator" xlink:label="srt_NameOfMajorCustomerDomain_2" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MajorCustomersAxis" xlink:type="locator" xlink:label="srt_MajorCustomersAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_SalesRevenueNetMember" xlink:type="locator" xlink:label="us-gaap_SalesRevenueNetMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskBenchmarkDomain" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskBenchmarkDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskByBenchmarkAxis" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskByBenchmarkAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_CustomerConcentrationRiskMember" xlink:type="locator" xlink:label="us-gaap_CustomerConcentrationRiskMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskTypeDomain" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskTypeDomain" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskByTypeAxis" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskByTypeAxis" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskTable" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskTable" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_ConcentrationRiskLineItems" xlink:type="locator" xlink:label="us-gaap_ConcentrationRiskLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="us-gaap_ConcentrationRiskLineItems" xlink:to="us-gaap_ConcentrationRiskTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ConcentrationRiskTable" xlink:to="us-gaap_ConcentrationRiskByTypeAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_ConcentrationRiskByTypeAxis" xlink:to="us-gaap_ConcentrationRiskTypeDomain" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ConcentrationRiskTypeDomain" xlink:to="us-gaap_CustomerConcentrationRiskMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ConcentrationRiskTable" xlink:to="us-gaap_ConcentrationRiskByBenchmarkAxis" order="10140.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="us-gaap_ConcentrationRiskByBenchmarkAxis" xlink:to="us-gaap_ConcentrationRiskBenchmarkDomain" order="10170.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ConcentrationRiskBenchmarkDomain" xlink:to="us-gaap_SalesRevenueNetMember" order="10190.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="us-gaap_ConcentrationRiskTable" xlink:to="srt_MajorCustomersAxis" order="10240.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="srt_MajorCustomersAxis" xlink:to="srt_NameOfMajorCustomerDomain_2" order="10250.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="srt_MajorCustomersAxis" xlink:to="srt_NameOfMajorCustomerDomain" order="10270.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_NameOfMajorCustomerDomain" xlink:to="onto_SKHynixIncMember" order="10290.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_NameOfMajorCustomerDomain" xlink:to="onto_TaiwanSemiconductorManufacturingCompanyLimitedMember" order="10310.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_NameOfMajorCustomerDomain" xlink:to="onto_SamsungSemiconductorMember" order="10330.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="us-gaap_ConcentrationRiskLineItems" xlink:to="us-gaap_ConcentrationRiskPercentage1" order="10360.00" priority="2" use="optional" />
  </link:definitionLink>
  <link:roleRef roleURI="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureShareRepurchaseAuthorizationTextualDetails" xlink:href="onto-20220702.xsd#Role_DisclosureShareRepurchaseAuthorizationTextualDetails" xlink:type="simple" />
  <link:definitionLink xlink:type="extended" xlink:role="http://www.ontoinnovation.com/20220702/taxonomy/role/DisclosureShareRepurchaseAuthorizationTextualDetails">
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockRepurchaseProgramRemainingAuthorizedRepurchaseAmount1" xlink:type="locator" xlink:label="us-gaap_StockRepurchaseProgramRemainingAuthorizedRepurchaseAmount1" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockRepurchasedAndRetiredDuringPeriodShares" xlink:type="locator" xlink:label="us-gaap_StockRepurchasedAndRetiredDuringPeriodShares" />
    <link:loc xlink:href="https://xbrl.fasb.org/us-gaap/2022/elts/us-gaap-2022.xsd#us-gaap_StockRepurchaseProgramAuthorizedAmount1" xlink:type="locator" xlink:label="us-gaap_StockRepurchaseProgramAuthorizedAmount1" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MinimumMember" xlink:type="locator" xlink:label="srt_MinimumMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_MaximumMember" xlink:type="locator" xlink:label="srt_MaximumMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeMember" xlink:type="locator" xlink:label="srt_RangeMember" />
    <link:loc xlink:href="https://xbrl.fasb.org/srt/2022/elts/srt-2022.xsd#srt_RangeAxis" xlink:type="locator" xlink:label="srt_RangeAxis" />
    <link:loc xlink:href="onto-20220702.xsd#onto_SharesRepurchaseAuthorizationTable" xlink:type="locator" xlink:label="onto_SharesRepurchaseAuthorizationTable" />
    <link:loc xlink:href="onto-20220702.xsd#onto_SharesRepurchaseAuthorizationLineItems" xlink:type="locator" xlink:label="onto_SharesRepurchaseAuthorizationLineItems" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="onto_SharesRepurchaseAuthorizationLineItems" xlink:to="onto_SharesRepurchaseAuthorizationTable" order="10010.00" priority="2" use="optional" xbrldt:contextElement="segment" xbrldt:closed="true" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="onto_SharesRepurchaseAuthorizationTable" xlink:to="srt_RangeAxis" order="10040.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="srt_RangeAxis" xlink:to="srt_RangeMember" order="10070.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_RangeMember" xlink:to="srt_MaximumMember" order="10090.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="srt_RangeMember" xlink:to="srt_MinimumMember" order="10110.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_SharesRepurchaseAuthorizationLineItems" xlink:to="us-gaap_StockRepurchaseProgramAuthorizedAmount1" order="10140.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_SharesRepurchaseAuthorizationLineItems" xlink:to="us-gaap_StockRepurchasedAndRetiredDuringPeriodShares" order="10180.00" priority="2" use="optional" />
    <link:definitionArc xlink:type="arc" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="onto_SharesRepurchaseAuthorizationLineItems" xlink:to="us-gaap_StockRepurchaseProgramRemainingAuthorizedRepurchaseAmount1" order="10220.00" priority="2" use="optional" />
  </link:definitionLink>
</link:linkbase>
