EpiOcular’sstrongcorrelationtohistoricalinvivodatasetsandexceptionalreproducibilitymakeitanidealmodelfortheocularsafetytestingofrawingredientsandfinalformulations. TheEpiOcularEyeIrritationTest(EIT)isaninvitroalternativetoanimaltestingandisavalidated,OECDtestguideline(TG492). EpiOcularhasbeenusedformanyyearsbyindustryasanon-animal,invitroalternativetodetermineDraizescores(forwatersolubletestarticles)andtoassessthemildness/ultra-mildness(sub-Draize)ofmaterialscontactingtheeyes.
Technology:
EpiOculareyeirritationassayshavebeenincorporatedintotoxicologylaboratoriesacrosstheworld.Withstraightforwardprotocolsandsimpleyethighlypredictiveendpoints,EpiOcularassaysareroutinelyutilizedthroughoutthecosmetic,personalcare,chemicalandpharmaceuticalindustries.Developedmorethan20yearsago,EpiOcularhasbeenusedfortestingthousandsofmaterials.Whetherscientistsneedariskassessmentassayfortheirritationpotentialofanoculardeliveredmaterialorthehazardassessmentofchemicalbeingtransportedacrosstheworld,EpiOcularisthetestsystemofchoicetomeetyouroculartestingneeds.
Applications:
OcularIrritationTests
EpiOcularEyeIrritationTest
TheEpiOcularEITisaninvitrohazardassessmentassaythathastheABIlitytodifferentiatematerialsthatareocularnon-irritantsfrommaterialsthatarenotocularirritants.TheEITisapplicabletoabroadrangeoforganicandinorganicchemicals,andisacceptedasOECDTG492.
EpiOcularEyeIrritationTestProtocol
WatchavideobyJoveonEpiOcularEIT
EpiOcularSubDraizeUltra-MildnessTest
TheEpiOcularUltra-mildnessTestisaninvitroriskassessmentassayroutinelyutilizedformildmaterialsforwhichtheDraizetestisinsensitive.ThishighlyreproducIBLeassayallowsforquantifiablediscriminationamongmild,milderandmildestproductformulations.
EpiOcularSubDraizeProtocol
EpiOcularMTTET-50(Neat)
Thisinvitroriskassessmentassayisbasedontheeffectivetimeatwhichamaterial(appliedneat)causesa50%reductionintissueviability(ET-50).BasedontheET-50,thetestarticleiscategorizedintooneof4classificationsrangingfromnon-irritatingtosevere/extremewhichcorrespondtogroupingsofRabbitDraizeEyeScores(MMAS).
EpiOcularMTTET-50NeatProtocol
EpiOcularMTTET-50(Dilution)
SimilartotheMTTET-50Neatmethod,thisinvitroriskassessmentassayisbasedontheeffectivetimeatwhichamaterial(dilutedto20%inwater)causesa50%reductionintissueviability(ET-50).BasedontheET-50,thetestarticleiscategorizedintooneof4classificationsrangingfromnon-irritatingtosevere/extremewhichcorrespondtogroupingsofRabbitDraizeEyeScores(MMAS).Thismethodisrecommendedforsurfactantbasedsolutionsandisapplicabletowater-solublematerialswithaspecificgravityof>0.95.
EpiOcularMTTET-50DilutionProtocol
BrowseourreferencelibrarytoseehowourEpiOculartissuehasbeenusedintheseareasofstudy.
TechSpecs:
Tissue:
Kit:StandardEpiOcular™kit(OCL-200)consistsof24tissues,eachtissue9mmindiameter.(Tissue“kits”containtissues,asmallamountofculturemedium,andplasticware;contactMatTekforspecifickitcontents.)
Substrate:Non-coated,9mmIDsinglewelltissuecultureplateinsertsareused(e.g.MillicellPCF,Nuncpolycarbonatesinglewelltissuecultureplateinserts).
Culture:Atairliquidinterface.
Histology:5-8celllayers(basallayertoapicalsurface).
Cornification:None.
LotNumbers:Tissuelotsproducedbyeachtechnicianforeachweekareassignedaspecificlotnumber.Typically,therearemultiplelotnumbersforanygivenweek’stissueproduction.Aletterofthealphabetisappendedtotheendofthelotnumbertodifferentiatebetweenindividualkitswithinagivenlotoftissues.Alltissuekitswithinalotareidenticalinregardstocells,medium,handling,cultureconditions,etc.
Shipment:At4°Conmedium-supplemented,agarosegelsin24-wellplate.
Shipmentday:EveryMonday.ShipmentonThursdayalsopossibleuponspecialrequest.
Delivery:TuesdaymorningviaFedExpriorityservice(US).OutsideUS:Tuesday-Thursdaydependingonlocation.
Shelflife:Includingtimeintransit,tissuesmaybestoredat4°Cforupto4dayspriortouse.However,extendedstorageperiodsarenotrecommendedunlessabsolutelynecessary.Inaddition,thebestreproducibilitywillbeobtainediftissuesareusedconsistentlyonthesameday,e.g.Tuesdayafternoonorfollowingovernightstorageat4°C(Wednesdaymorning).
Alternativetissues:
OCL-200-ABF:Antibiotic-freetissue.Forlast3daysofculture,gentamicinisomittedfromculturemedium.Customerreceives200mlofOCL-200-ASY-AFABmedium.
OCL-200-AFF:Anti-fungal-freetissue.Forlast3daysofculture,Amphotericinisomittedfromculturemedium.Customerreceives200mlofOCL-200-ASY-AFABmedium.
OCL-200-AFAB:Antibiotic,anti-fungal-freetissue.Forlast3daysofculture,gentamicinandAmphotericinareomittedfromculturemedium.Customerreceives200mlofOCL-200-ASY-AFABmedium.
OCL-200-EACH:SingleEpiOculartissueinsert.
OCL-200-FRZN-EA:SingleOCL-200tissuethathasbeensubjectedto3freeze/thawcycles.DesignedtoserveascontrolforMTTauto-reductiontest.
OCL-212:Halfkit(12culturesinsteadofstandard24).
OCL-606:SameasOCL-200except6tissuesperkit,eachtissueculturedinalargercellcultureinsert(22mmdiameter).
Cells:
Type:Normalhumanepidermalkeratinocytes(NHEK).
Geneticmake-up:Singledonor.
Derivedfrom:Neonatal-foreskintissue.
Screenedfor:HIV,Hepatitis-B,Hepatitis-C,mycoplasma.
Medium:
Basemedium:Dulbecco’sModifiedEagle’sMedium(DMEM).
Growthfactors/hormones:Epidermalgrowthfactor,insulin,hydrocortisoneandotherproprietarystimulatorsofnon-keratinizingepithelialdifferentiation.
Serum:None.
Antibiotics:Gentamicin5µg/ml(10%ofnormalgentamicinlevel).
Anti-fungalagent:AmphotericinB0.25µg/ml.
pHIndicator:Phenolred.
Alternatives:Phenolred-free,antibiotic-free,anti-fungal-free,orhydrocortisone-freemediumandtissueareavailable.Agentsareremovedatleast3dayspriortoshipment.
Maintenancemedium:MostassayswithEpiOcular™areperformedwithin24hours.However,forlongerexperiments,thetissuecanbemaintainedinOCL-200-MM(identicaltoEpiOcular™assaymedium).
QualityControlandSterility:
Visualinspection:Alltissuesarevisuallyinspectedandifphysicalimperfectionsarenoted,tissuesarerejectedforshipment.
End-usetesting:Tissuesareexposedto0.3%TritonX-100for5,20,and60minutes.Thetimeofexposurerequiredtoreducethetissueviability(ET-50)usingtheMTTassayisdetermined(SeeMatTekEpiOcularTMET-50protocol)foreachlotoftissue.ET-50’smustfallwithintherangeofthe1996EpiOcular™database:12.2-37.5minutes.ET-50’sincustomers’labsareoftenverysimilartoMatTekresults.
Sterility:Allmediausedthroughouttheproductionprocessischeckedforsterility.Maintenancemediumisincubatedwithandwithoutantibioticsfor1weekandcheckedforsterility.Theagarosegelfromthe24-wellplateusedforshippingisalsoincubatedfor1weekandcheckedforanysignofcontamination.
Screeningforpathogens:AllcellsarescreenedandarenegativeforHIV,hepatitisBandhepatitisCusingPCR.However,noknowntestmethodcanoffercompleteassurancethatthecellsarepathogenfree.Thus,theseproductsandallhumanderivedproductsshouldbehandledatBSL-2levels(biosafetylevel2)orhigherasrecommendedintheCDC-NIHmanual,“BiosafetyinmicroBIOLOGicalandbiomedicallaboratories,”1998.Forfurtherassistance,pleasecontactyoursiteSafetyOfficerorMatTektechnicalservice.
Notificationoflotfailure:IfatissuelotfailsourQCorsterilitytesting,thecustomerwillbenotifiedandthetissueswillbereplacedwithoutcharge.BecauseourQCandsterilitytestingisdonepost-shipment,notificationwillbemadeassoonaspossible(Undernormalcircumstances,ET-50failureswillbenotifiedbyWednesday5pm;sterilityfailureswillbenotifiedwithin8daysofshipment).
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