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AntibodiesOnline/Fibronectin ELISA Kit/ABIN491751/96 tests188bio精品生物—专注于实验室精品爆款的电商平台 - 蚂蚁淘旗下精选188款生物医学科研用品
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AntibodiesOnline/Fibronectin ELISA Kit/ABIN491751/96 tests
Antigen
Fibronectin
  • FN
  • cig
  • fibronectin
  • finc
  • lets
  • msf
  • hypotheticalprotein
  • fibronectin1
  • fibronectin
  • fn1
Reactivity
Human
Alternatives
Kitswithalternativereactivityto:
29Human
25Mouse(Murine)
21Rat(Rattus)
9Cow(Bovine)
8Pig(Porcine)
2Monkey
2Rabbit
2Wildboar(Susscrofa)
1Chicken
1Dog(Canine)
1Goat
1GuineaPig
1Horse(Equine)
1Sheep(Ovine)
MethodType
SandwichELISA
DetectionRange
0.27-200ng/mL
MinimumDetectionLimit
0.27ng/mL
Application
ELISA
Options
Bulkdiscount
Supplier
SupplierProductNo.
SampleTypeSerum,Plasma,TissueHomogenate,CellLysate,CellCultureSupernatant,BiologicalFluids
AnalyticalMethodQuantitative
DetectionMethodChemiluminescent
Specificity

ThisassayhashighsensitivityandexcellentspecificityfordetectionofFibronectin(FN).
Nosignificantcross-reactivityorinterferencebetweenFibronectin(FN)andanalogueswasobserved.

Cross-Reactivity(Details)Nosignificantcross-reactivityorinterferencebetweenFibronectin(FN)andanalogueswasobserved.
Sensitivity0.12ng/mL
Components
  • Pre-coated,readytouse96-wellstripplate
  • Platesealerfor96wells
  • StandardDiluent
  • AssayDiluentA
  • AssayDiluentB
  • StopSolution
  • Standard
  • DetectionReagentA
  • DetectionReagentB
  • TMBSubstrate
  • WashBuffer(30×concentrate)
  • Instructionmanual
Materialnotincluded
  • Luminometercapableofreading96-wellmicroplateswiththefollowingparameters:lagtime30.0secs,readtime1.0sec/well.
  • Precisionsingleormulti-channelpipettesandpipettetipswithdisposabletips.
  • EppendorfTubesfordilutingsamples.
  • Deionizedordistilledwater.
  • Absorbentpaperforblottingthemicrotiterplate.
  • ContainerforWashSolution
AlternativeNameFibronectin(FN)
UniProtP02751
ApplicationNotes
  • Limitedbythecurrentconditionandscientifictechnology,wecannotcompletelyconductthecomprehensiveidentificationandanalysisontherawmaterialprovidedbysuppliers.Sotheremightbesomequalitativeandtechnicalriskstousethekit.
  • Thefinalexperimentalresultswillbecloselyrelatedtovalidityoftheproducts,operationskillsoftheendusersandtheexperimentalenvironments.Pleasemakesurethatsufficientsamplesareavailable.
  • Kitsfromdifferentbatchesmaybealittledifferentindetectionrange,sensitivityandcolordevelopingtime.
  • Donotmixorsubstitutereagentsfromonekitlottoanother.Useonlythereagentssuppliedbymanufacturer.
  • Protectallreagentsfromstronglightduringstorageandincubation.Allthebottlecapsofreagentsshouldbecoveredtightlytopreventtheevaporationandcontaminationofmicroorganism.
  • Theremaybesomefoggysubstanceinthewellswhentheplateisopenedatthefirsttime.Itwillnothaveanyeffectonthefinalassayresults.Donotremovemicrotiterplatefromthestoragebaguntilneeded.
  • Wrongoperationsduringthereagentspreparationandloading,aswellasincorrectparametersettingfortheplatereadermayleadtoincorrectresults.Amicroplateplatereaderwithabandwidthof10nmorlessandanopticaldensityrangeof0-3O.D.orgreaterat450±10nmwavelengthisacceptableforuseinabsorbancemeasurement.Pleasereadtheinstructioncarefullyandadjusttheinstrumentpriortotheexperiment.
  • Eventhesameoperatormightgetdifferentresultsintwoseparateexperiments.Inordertogetbetterreproducibleresults,theoperationofeverystepintheassayshouldbecontrolled.Furthermore,apreliminaryexperimentbeforeassayforeachbatchisrecommended.
  • EachkithasbeenstrictlypassedQ.Ctest.However,resultsfromendusersmightbeinconsistentwithourin-housedataduetosomeunexpectedtransportationconditionsordifferentlabequipments.Intra-assayvarianceamongkitsfromdifferentbatchesmightarisefromabovefactors,too.
  • Kitsfromdifferentmanufacturersforthesameitemmightproducedifferentresults,sincewehavenotcomparedourproductswithothermanufacturers.
Comment

Informationonstandardmaterial:
Thestandardmightberecombinantproteinornaturalprotein,thatwilldependonthespecifickit.Moreover,theexpressionsystemisE.colioryeastormammalcell.Thereis0.05%proclin300inthestandardaspreservative.

Informationonreagents:
Thestopsolutionusedinthekitissulfuricacidwithconcentrationof1mol/L.AndthewashsolutionisTBS.Thestandarddiluentcontains0.02%sodiumazide,assaydiluentAandassaydiluentBcontain0.01%sodiumazide.SomekitscancontainisBSAinthem.

Informationonantibodies:
Theprovidedantibodiesandtheirhostvaryindifferentkits.

SampleVolume100μL
AssayTime3h
PlatePre-coated
ProtocolThemicroplateprovidedinthiskithasbeenpre-coatedwithanantibodyspecifictoFibronectin(FN).Standardsorsamplesarethenaddedtotheappropriatemicroplatewellswithabiotin-conjugatedantibodyspecifictoFibronectin(FN).Next,AvidinconjugatedtoHorseradishPeroxidase(HRP)isaddedtoeachmicroplatewellandincubated.ThenthemixtureofsubstrateAandBisaddedtogenerateglowlightemissionkinetics.Uponplatedevelopment,theintensityoftheemittedlightisproportionaltotheFibronectin(FN)levelinthesampleorstandard.,
ReagentPreparation
  • Bringallkitcomponentsandsamplestoroomtemperature(18-25°C)beforeuse.
  • Standard:ReconstitutetheStandardwiththeStandardDiluent,keepatroomtemperaturefor10minandshakegently(nottofoam).PreparetubescontainingStandardDiluenttoproduceadoubledilutionseries.
  • AssayDiluentAandAssayDiluentB:Dilute6mLofAssayDiluentAorBConcentrate(2×)with6mLofdeionizedordistilledwatertoprepare12mLofAssayDiluentAorB.(Infact,morethan6mLAssayDiluentAandAssayDiluentBarecontainedinthebottles.Therefore,ineverytest,pleasepreciselypipetterequiredamountofDiluentandmakedoubledilutioninanewcontainer.Thepreparedworkingdilutioncannotbefrozen.)
  • DetectionReagentAandDetectionReagentB:BrieflyspinorcentrifugethestockDetectionAandDetectionBbeforeuse.DilutetotheworkingconcentrationwithworkingAssayDiluentAorB,respectively(1:100).
  • WashSolution:Dilute20mLofWashSolutionconcentrate(30×)with580mLofdeionizedordistilledwatertoprepare600mLofWashSolution(1×).
  • SubstrateworkingSolution:MixthesubstrateAandBbytheratioof99:1tomakethesubstrateworkingsolution.Mixthoroughly.Forexample,prepare1,000μLSubstrateworkingSolutionwith990μLSubstrateA+10μLSubstrateB.
Note:
  • Makingserialdilutioninthewellsdirectlyisnotpermitted.
  • Preparestandardwithin15minutesbeforeassay.Pleasedonotdissolvethereagentsat37°Cdirectly.
  • PleasecarefullyreconstituteStandardsorworkingDetectionReagentAandBaccordingtotheinstruction,andavoidfoamingandmixgentlyuntilthecrystalsarecompletelydissolved.Tominimizeimprecisioncausedbypipetting,usesmallvolumesandensurethatpipettorsarecalibrated.Itisrecommendedtosuckmorethan10μLforoncepipetting.
  • ThereconstitutedStandards,DetectionReagentAandDetectionReagentBcanbeusedonlyonce.
  • PrepareSubstrateworkingSolutionwithin15minutesbeforeassay.
  • IfcrystalshaveformedintheWashSolutionconcentrate(30×),warmtoroomtemperatureandmixgentlyuntilthecrystalsarecompletelydissolved.
  • Contaminatedwaterorcontainerforreagentpreparationwillinfluencethedetectionresult.
SampleCollectionSerum:Allowsamplestoclotfortwohoursatroomtemperatureorovernightat4°Cbeforecentrifugationfor20minutesatapproximately1000×g.Assayimmediatelyorstoresamplesinaliquotat-20°Cor-80°C.Avoidrepeatedfreeze/thawcycles.

Plasma:CollectplasmausingEDTAorheparinasananticoagulant.Centrifugesamplesfor15minutesat1000×gwithin30minutesofcollection.Removeplasmaandassayimmediatelyorstoresamplesinaliquotat-20°Cor-80°C.Avoidrepeatedfreeze/thawcycles.

BiologicalFluids:Centrifugesamplesfor20minutesat1000×g.Removeparticulatesandassayimmediatelyorstoresamplesinaliquotat-20°Cor-80°Cforlateruse.Avoidrepeatedfreeze/thawcycles.
SamplePreparation
  • Weareonlyresponsibleforthekititself,butnotforthesamplesconsumedduringtheassay.Theusershouldcalculatethepossibleamountofthesamplesusedinthewholetest.Pleasereservesufficientsamplesinadvance.
  • Pleasepredicttheconcentrationbeforeassaying.Ifvaluesforthesearenotwithintherangeofthestandardcurve,usersmustdeterminetheoptimalsampledilutionsfortheirparticularexperiments.Sampleshouldbedilutedby0.01mol/LPBS(PH=7.0-7.2).
  • Ifthesamplesarenotindicatedinthemanual,apreliminaryexperimenttodeterminethevalidityofthekitisnecessary.
  • TissueorcellextractionsamplespreparedbychemicallysisbuffermaycauseunexpectedELISAresultsduetotheimpactsfromcertainchemicals.
  • Duetothepossibilityofmismatchingbetweenantigenfromotheroriginandantibodyusedinourkits(e.g.antibodytargetsconformationalepitoperatherthanlinearepitope),somenativeorrecombinantproteinsfromothermanufacturersmaynotberecognizedbyourproducts.
  • Influencedbythefactorsincludingcellviability,cellnumberorsamplingtime,samplesfromcellculturesupernatantmaynotbedetectedbythekit.
  • Freshsampleswithoutlongtimestorageisrecommendedforthetest.Otherwise,proteindegradationanddenaturalizationmayoccurinthosesamplesandfinallyleadtowrongresults.
AssayProcedure
  1. Prepareallreagents,samplesandstandards,
  2. Add100μLstandardorsampletoeachwell.Incubate2hoursat37 °C,
  3. Aspirateandadd100μLpreparedDetectionReagentA.Incubate1hourat37 °C,
  4. Aspirateandwash3times,
  5. Add100μLpreparedDetectionReagentB.Incubate30 minutesat37 °C,
  6. Aspirateandwash5times,
  7. Add100μLSubstrateSolution.Incubate10 minutesat37 °C,
  8. ReadRLUvalueimmediately.
CalculationofResults

Averagetheduplicatereadingsforeachstandard,control,andsamplesandsubtracttheaveragezerostandardrelativelightunit(RLU).Createastandardcurveonlog-loggraphpaper,withCNTFconcentrationonthey-axisandtheRLUvalueonthex-axis.Drawthebestfitstraightlinethroughthestandardpointsanditcanbedeterminedbyregressionanalysis.Usingsomeplotsoftware,suchascurveexpert1.30,isalsorecommended.Ifsampleshavebeendiluted,theconcentrationreadfromthestandardcurvemustbemultipliedbythedilutionfactor.

AssayPrecision

Intra-assayPrecision(Precisionwithinanassay):3sampleswithlow,middleandhighlevelFibronectin(FN)weretested20timesononeplate,respectively.
Inter-assayPrecision(Precisionbetweenassays):3sampleswithlow,middleandhighlevelFibronectin(FN)weretestedon3differentplates,8replicatesineachplate.
CV(%)=SD/meanX100
Intra-Assay:CV<10%
Inter-Assay:CV<12%

RestrictionsForResearchUseonly
PrecautionofUseTheStopSolutionsuggestedforusewiththiskitisanacidsolution.Weareye,hand,face,andclothingprotectionwhenusingthismaterial.
HandlingAdvice

Thestabilityofkitisdeterminedbythelossrateofactivity.Thelossrateofthiskitislessthan5 %withintheexpirationdateunderappropriatestoragecondition.
Tominimizeextrainfluenceontheperformance,operationproceduresandlabconditions,especiallyroomtemperature,airhumidity,incubatortemperatureshouldbestrictlycontrolled.Itisalsostronglysuggestedthatthewholeassayisperformedbythesameoperatorfromthebeginningtotheend.

Storage4°C
StorageComment
  • Forunopenedkit:Allthereagentsshouldbekeptaccordingtothelabelsonvials.TheStandard,DetectionReagentA,DetectionReagentBandthe96-wellstripplateshouldbestoredat-20°Cuponreceiptwhiletheothersshouldbeat4°C.
  • Foropenedkit:Whenthekitisopened,theremainingreagentsstillneedtobestoredaccordingtotheabovestoragecondition.Besides,pleasereturntheunusedwellstothefoilpouchcontainingthedesiccantpack,andresealalongentireedgeofzip-seal.
    Note:Itishighlyrecommendedtousetheremainingreagentswithin1monthprovidedthisiswithintheexpirationdateofthekit.
  • ForELISAkit,1daystorageat37°Ccanbeconsideredas2monthsat4°C,whichmeans3daysat37°Cequaling6monthsat4°C.
ExpiryDate6months