EpiVaginalprovidesaninvitroresearchtoolbothforfeminineproductsafetytestingandforthestudyofHIV-1andothersexuallytransmitteddiseases.Itstissuestructureandcellularphysiologycloselyparallelinvivovaginalepithelialtissue,makingitidealfortoxicologyandefficacytesting,aswellasatoolforBIOLOGicalresearch.
Technology:
EpiVaginaltissuesareculturedfromnormal,human-derivedvaginalepithelialanddendriticcells.Itshighly-differentiatedstructureparallelsinvivotissueandisidealfortoxicitystudiesoffemininehygiene,vaginalcare,andmicrobicideproducts.FourtypesofEpiVaginalareoffered:
1)VEC-100(Figure1):AnepithelialtissuecontainingepithelialVECcells,
2)VLC-100(Figure1):ApatentedepithelialtissuecontainingepithelialVECandimmuno-competentdendriticcells,
3)VEC-100-FT(Figure2):AfullthicknessversionofVEC-100whichincludesVECepithelialcellsandafibroblast-containinglaminapropria,and
4)VLC-100-FT(Figure2):Apatentedimmuno-competentversionoftheVEC-100-FTwhichincludesdendriticcells.
Figure1:H&Estainedhistological(formalinfixed)cross-sectionsofA)VEC-100invitroreconstructedepithelialtissuemodelscontainingnormalhumanVECcells,andB)vaginalexplanttissue.Bothinvitroandinvivotissuesshownucleatedbasalandsuprabasalcelllayersfollowedbylayersinwhichnucleiarelostandcellsarefilledwithglycogen.
Figure2:H&Estainedhistological(formalinfixed)crosssectionsoffull-thicknessEpiVaginaltissues.TheVEC-100-FTtissuesconsistofVECepithelialcellsculturedatopalaminapropria(LP)-likecollagenmatrixthatcontainsfibroblasts.TheVLC-100-FTconsistsofVECepithelialanddendriticcellsintheepitheliallayersandcontainsbothfibroblastsanddendriticcellsintheLP.
Figure3:Formalinfixedcross-sectionsoftheVEC-100tissueimmuno-stainedforcytokeratins,CK13,CK14,&CK18.BasalcellsarestainedbyCK14andsupra-basalcellsbyCK13;nocellsarestainedbyCK18.
Figure4:ViABIlity(A)andCytokineRelease(B)offull-thicknessvaginalecto-cervical(VEC-100-FT)tissuemodelfollowingexposure(18hr)toformulationscontainingthecommonspermicidenonoxynol-9(N9):a)PBScontrolandb)KYjelly(KY,0%N9),c)KYN9(2%),andd)N9(0.1%).Asviabilityofthetissuedecreases,theIL-1βincreases.
Figure5:HIV-1infectionofVLC-100tissuefollowingtopicalexposure(24hr)to60,000CPMofHIV-1ADAorNL4-3virus.Afterexposure,tissueswerewashed3Xandthenculturedforanadditional24hr.CellularDNAwasthenextractedandHIV-1transcriptsweredetectedusingHIV-1gagspecificprimerpairs.
Figure6:RTPCRshowingToll-likeReceptor(TLR)expressionfor:A)organotypicvaginal-ectocervical(VEC)tissuemodelandB)Humancervicaltissueexplant.
Applications:
MicrobicideTesting,STDInfectionResearch,FeminineHygiene,Inflammation,DrugDelivery
EpiVaginalprovidesaninvitroresearchtoolbothforfeminineproductsafetytestingandforthestudyofHIV-1andothersexuallytransmitteddiseases.Itstissuestructureandcellularphysiologycloselyparallelinvivovaginalepithelialtissue,makingitidealfortoxicologyandefficacytesting,aswellasatoolforbiologicalresearch.BrowseourreferencelibrarytoseehowresearchershaveusedourEpiVaginaltissueintheseareasofstudy.
TechSpecs:
Tissue
Kits:TheEpiVaginalkitVEC-100contains24tissues,VEC-112contains12tissuesandVEC-606contains6tissues.(Tissue“kits”containtissues,asmallamountofculturemedium,andplasticware;contactMatTekforspecifickitcontents.)
Substrate:Singlewelltissuecultureplateinsertsareused.
VEC-100:Nunc™singlewelltissuecultureplateinserts.PoreSize=0.4µm,InnerDiameter=0.80cm,SurfaceArea=0.5cm2.
VEC-606:Polycarbonatesinglewelltissuecultureplateinserts,Nunc™.PoreSize=0.4µm,InnerDiameter=2.5cm,SurfaceArea=4.2cm2.
Culture:Atairliquidinterface.
Histology:VEC-100:10-16celllayersofnon-cornifiedtissue.
LotNumbers:Tissuelotsproducedbyeachtechnicianforeachweekareassignedaspecificlotnumber.Typically,therearemultiplelotnumbersforanygivenweek’stissueproduction.Aletterofthealphabetisappendedtotheendofthelotnumbertodifferentiatebetweenindividualkitswithinagivenlotoftissues.Alltissuekitswithinalotareidenticalwithregardtocells,medium,handling,cultureconditions,etc.
Shipment:At4°Conmedium-supplemented,agarosegels.
Shipmentday:Monday.
Delivery:TuesdaymorningviaFedExpriorityservice(US).OutsideUS:Tuesday-Thursdaydependingonlocation.
Shelflife:Includingtimeintransit,tissuesmaybestoredat4°Cforupto4dayspriortouse.However,extendedstorageperiodsarenotrecommendedunlessabsolutelynecessary.Inaddition,thebestreproducibilitywillbeobtainediftissuesareusedconsistentlyonthesameday,e.g.Wednesdaymorningfollowingovernightstorageat4°C.
Lengthofexperiments:Culturescanbecontinuedforatleast1weekwithgoodretentionofnormalmorphology.Culturesmustbefedeveryotherdaywith5.0mlofmaintenancemedium(VEC-100-MM).Cellcultureinsertsareplacedatopculturestands(partnumberMEL-STND)orwashers(partnumberEPI-WSHR)in6-wellplatestoallowtheuseof5.0ml.
Alternativetissues:
VEC-100-FT:SimilartoVEC-100excepttissuegrownonacollagen/vaginalfibroblastsubstrate(laminapropria).
VEC-112-FT:SimilartoVEC-100-FTexcept12tissues.
VLC-100:SimilartoVEC-100excepttissuescontainLangerhanscells.
VLC-112:SimilartoVLC-100except12tissues.
VLC-100-FT:SimilartoVLC-100excepttissuegrownonacollagen/vaginalfibroblastsubstrate(laminapropria).
VLC-112-FT:SimilartoVLC-100-FTexcept12tissues.
Cells
Type:Normalhumanectocervicalcells(NHEC)aredifferentiatedintotissueswithanon-cornified,vaginal-ectocervicalphenotype(partnumberVEC-100).
Geneticmake-up:Singledonor.
Derivedfrom:adulthumanectocervicaltissue.
Screenedfor:HIV,Hepatitis-B,Hepatitis-C,mycoplasma,bacteria,yeast,fungi.
Medium
Basemedium:Dulbecco’sModifiedEagle’sMedium(DMEM)andF12Medium.
Growthfactors/hormones:Epidermalgrowthfactorandotherproprietaryfactors.
Serum:None.
Antibiotics:Gentamicin5µg/ml(10%ofnormalgentamicinlevel).
Anti-fungalagent:AmphotericinB0.25µg/ml.
pHIndicator:Phenolred.
Otheradditives:Proprietary.
Alternatives:Phenolred-free,antibiotic-free,anti-fungal-freemediumandtissueareavailable.Agentsareremoved3dayspriortoshipment.
Maintenancemedium:VEC-100-MM
QualityControlandSterility
Visualinspection:Alltissuesarevisuallyinspectedandifphysicalimperfectionsarenoted,tissuesarerejectedforshipment.
End-usetesting:TissueviabilityofeachlotisdeterminedbyanMTTassay.Tissuesareexposedto1.0%TritonX-100for30,60,and120minutes.Thetimeofexposurerequiredtoreducethetissueviability(ET-50)usingtheMTTassayisdetermined.Thetimeofexposureneededtoreducetheviabilityto50%(ET-50)mustbe0.92–1.67hours.AsummaryoftheQCtestingresultswillbeprovidedtothecustomerforreferenceuponrequest.(ForTEERmeasurements,MatTekhassuccessfullyusedtheWPIEndohmSystem.)
Sterility:Allmediausedthroughouttheproductionprocessischeckedforsterility.Maintenancemediumisincubatedwithandwithoutantibioticsfor1weekandcheckedforsterility.Theagarosegelfromthe24-wellplateusedforshippingisalsoincubatedfor1weekandcheckedforanysignofcontamination.
Screeningforpathogens:AllcellsarescreenedandarenegativeforHIV,hepatitisBandhepatitisCusingPCR.However,noknowntestmethodcanoffercompleteassurancethatthecellsarepathogenfree.Thus,theseproductsandallhumanderivedproductsshouldbehandledatBSL-2levels(biosafetylevel2)orhigherasrecommendedintheCDC-NIHmanual,“Biosafetyinmicrobiologicalandbiomedicallaboratories,”1998.Forfurtherassistance,pleasecontactyoursiteSafetyOfficerorMatTektechnicalservice.
Notificationoflotfailure:IfatissuelotfailsourQCorsterilitytesting,thecustomerwillbenotifiedandthetissueswillbereplacedwithoutchargewhenappropriate.BecausesomeofourQCandsterilitytestingisdonepost-shipment,notificationwillbemadeassoonaspossIBLe.(Undernormalcircumstances,ET-50failureswillbenotifiedbyThursday,5p.m.;sterilityfailureswillbenotifiedwithin8daysofshipment.)
