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IBL/IGF-1 direct (Rat, Mouse) ELISA/30117393/

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Kitsize12x8MethodELISAIncubationtime1x1h,1x30min,1x30minStandardrange0.5-18µg/L,AssayRange:0.029-180µg/LSpecimen/Volumes5µLserum,plasma/withoutextractionSubstrate/isotopeTMB450nmRegulatoryStatus:ForresearchuseonlyDetailsfor: IGF-1direct(Rat,Mouse)ELISAMouse/RatIGF-IELISAissuitedforIGF-Ideterminationinserumandplasmaofmiceandratshighsensitive:0.029ng/mlanalyticalsensitivity;requiredsamplevolumeisverysmallisfast:incubationtimeatotalof2hoursSingleStandardswith0.5,2.5,6,12,18ng/mlrecombinantIGF-IareprovidedintheKit2ControlSeraareprovidedforqualitycontroluseshighaffinityantibodiesagainstm/rIGF-IMicrotiterplatesareseparatelybreakapartIntendedUseMeasurementofIGF-Iinmouse/ratserumandplasma.INTRODUCTIONBesidedifferentcellculturemodelsandstudieswithhumanpatients,miceandratsaresuitablemodelorganismsforbasicresearchandpre-clinicalstudies.Thus,wedevelopedthistestsystemasatoolforIGF-Imeasurementsinmiceandratforusageinresearchandpreclinicalstudies.EvenifthecomparABIlityofmiceandhumansislimitedweoffersomebackgroundinformationonthehumanIGF-Isysteminthefollowingsection:Insulin-likegrowthfactors(IGF)IandIIplayapivotalroleinregulatingtheproliferationanddifferentiationofmanycelltypes.IGF-IisidenticalwithSomatomedinC(Sm-C)andhasamolecularweightof7649daltons.Itsmajorregulatorsaregrowthhormone(GH)andnutrition(6).Incontrasttomanyotherpeptidehormones,IGFsareavidlyboundtospecificbindingproteins(IGFBP).ThesevenIGFBPswhichareknownatpresenteitherbindIGF-IandIGF-IIwithsimilaraffinitiesorshowapreferenceforIGF-II.AmajorproblemofIGF-ImeasurementresultsfromtheinterferenceofIGFBPsintheassay.DirectdeterminationsinuntreatedserumsamplesgivefalsevaluesbecauseoftheextremelyslowdissociationoftheIGF-I/IGFBP-3complexesduringtheassayincubation.DependingontheratioIGF-ItoIGFBPinthesampleinterferencecomesup.Therefore,varioustechniqueswereappliedtophysicallyseparateIGF-Ifromitsbindingproteinsbeforemeasurement,including(a)sizeexclusionchromatographyunderacidicconditions,(b)solid-phaseextractionand(c)acid-ethanolextraction.Thesetechniques,however,areeitherinconvenientortime-consumingorgiveincompleteandnotreproducIBLerecoveries.Themostwidelyusedmethodistheacid-ethanolextractionwitharecoveryofonly70-80%ofIGFBP-boundIGF-Iasaresultofco-precipitation.Theabsoluteresultsofsuchanextractionarethereforefalselow.TheextractionremovestheIGFBPsonlyinsufficientlyandleadstoreductioninsensitivityoftheassayduetopredilutionofthesamplesbytheextractionprocedure.FurThermore,theremainingIGFBPmaystillinterfereintheassay.Inaddition,theacid-ethanolextractionisineffectiveinspecimensotherthanserumorplasma(e.g.cellculturemedia),inwhichdeterminationofIGF-IisalreadydifficultenoughduetothefactthatIGFBPsarefrequentlypresentatlargeexcess.Toavoidthesedifficulties,anuncomplicatedassaywasdeveloped,inwhichspecialsamplepreparationisnotrequiredbeforemeasurement.ForconcretedatapleaseconsulttheInstructionforUseinthedownloadboxontherightside.

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