CA-N426 | |
Name | EliteTMNADP/NADPHRatioAssayKit(RedFluorescence) |
Description | ThisElite™FluorimetricNADP/NADPHRatioAssayKitprovidesaconvenientmethodforsensitivedetectionofNADP,NADPHandtheirratio.TheenzymesinthesystemspecificallyrecognizeNADP/NADPHinanenzymerecyclingreactionthatsignificantlyincreasesdetectionsensitivity.Inaddition,thisassayhasverylowbackgroundsinceitisrunintheredvisIBLerangethatconsiderablyreducesthesampleinterference. |
Application | Theassaycanbeperformedinaconvenient96-wellor384-wellmicrotiter-plateformat.ItssignalcanbeeasilyreadbyeitherafluorescencemicroplatereaderatEx/Em=540/590nmoranabsorbancemicroplatereaderat~576nm. |
Size | 250assaysin96-wellplates |
Ex/Em | 540/590nm |
Detection | Fluorescencemicroplatereaderorabsorbancemicroplatereader |
Clickherefor DATASHEET | Clickherefor MSDS |
FrequentlyAskedQuestions
1.CanweusethisratioassaykittoalsodetectNADPorNADPHseparately?
Answer:Yes.YoucouldmeasuretheNADOHamountwithComponentD(NADPHExtractionSolution),andmeasuretheNADPamountwithNADPExtractionSolution.Butaccordingtoourexperience,theNADPconcentrationsincellsarealothigherthantheNADPHconcentrations.WesuggesttomeasuretheTotalNADP+NADPHamountandtheNADPamount,thentocalculatetheNADPHamountbysubtraction,whichgivesmoreaccurateresults.
2.WhyPBSisusedtodilutetheStandard?Sincethesamplesareinthelysisbuffer,doyouthinkIshouldusethelysisbuffertodilutethem?
Answer:PBSiscommonlyused.Peopleusuallyusethelysisbuffertolysethecells,thenusePBStodilutethesamples.ThepointisthatyoushouldkeepthesameconcentrationofthelysisbufferforthesamplesandtheStandards.
3.Ididacomparison,anditshowedthatthePBSbasedStandardcurvelookedbetterthanthelysisbufferbasedStandardcurve.Whatdoyouthinkofthis?
Answer:ThebubblesinthelysisbuffermaycausetheinconsistencyofthereADIngs.
4.WhenIdoduplicatesofStandards,thereadingvariesalotinthelowrange(0.2uMorlower).Whydoesthathappen?
Answer:Itisnormal.Therearemanyfactorsthatwouldcauseinconsistentreadings,i.e.bubblesinthewells,thevolumes....Sotrydoingmorereplicates.
5.Sometimesforthe2setsofStandardsIsetup,theylookgood(R2=0.99)individuallybutthecurveismessedup(R2=0.95)whenmerged.SowhichcurveshouldIusetoquantitatemydata?
Answer:You"llneedtousethecurvesetuponthesameplatewithyoursamples.
6.Wedon"thavefluorescencemicroplatereader,socanweuseabsorbancemicroplatereader?
Answer:Yes.Buttheabsorbancereadingshavemuchlowersensitivitycomparedtothefluorescencereadings.Tryusingtheratioof570to610toincreasethesensitivity.