Overview:
ProductName | NitricOxideDetectionKit |
Description | Colorimetricdetectionofnitrateandnitrite |
SpeciesReactivity | SpeciesIndependent |
Platform | Microplate |
SampleTypes | Buffer,Celllysates,Plasma,Saliva,Serum,TissueCultureMedia,Urine,Water |
DetectionMethod | ColorimetricAssay |
AssayType | IndirectQuantitativeAssay |
Utility | ColorimetricassayusedtoquantitativelymeasureNitrateandNitritepresentinavarietyofsamples. |
Sensitivity | 2.63µMintheNitriteand1.02µMintheTotalNitricOxide |
AssayRange | 3.125-200µM |
Precision | IntraAssayPrecision:ThreesampleswerefurtherdilutedinAssayBufferandruninreplicatesof20inanassay.ThemeanandprecisionofthecalculatedNitriteorTotalNOconcentrationswere:Sample1(NitriteConcentration)-45.1µM,4.4%CV,Sample1(TotalNOConcentration)-70.5µM,6.8%CVSample2(NitriteConcentration)-73.3µM,9.1%CV,Sample1(TotalNOConcentration)-107.4µM,4.4%CVSample3(NitriteConcentration)-132.7µM,1.3%CV,Sample1(TotalNOConcentration)-157.8µM,1.8%CVInterAssayPrecision:ThreesampleswerefurtherdilutedinAssayBufferandruninduplicatesintwentyassaysrunovermultipledaysbythreeoperators.ThemeanandprecisionofthecalculatedNitriteorTotalNOconcentrationswere:Sample1(NitriteConcentration)-44.1µM,3.1%CV,Sample1(TotalNOConcentration)-68.8µM,7.4%CVSample2(NitriteConcentration)-66.4µM,4.0%CV,Sample1(TotalNOConcentration)-112.1µM,5.7%CVSample3(NitriteConcentration)-126.7µM,6.3%CV,Sample1(TotalNOConcentration)-154.48µM,4.1%CV |
IncubationTime | 30minutes |
NumberofSamples | 88samplesinduplicate |
OtherResources | KitBooklet,MSDS |
Properties
StorageTemperature | 4ºC | ||||||||||||||||||||||||||||||
ShippingTemperature | BlueIce | ||||||||||||||||||||||||||||||
ProductType | DetectionKits | ||||||||||||||||||||||||||||||
AssayOverview | TheNitricOxideDetectionKitisdesignedtoquantitativelymeasureNitrateandNitritepresentinavarietyofsamples.NitricOxidecontentisderivedfromthesumofNitrate(-NO3)andNitrite(-NO2).BothNitrateandNitritestandardsareprovidedtogeneratestandardcurvesfortheassayandallsamplesshouldbereadofftheappropriatestandardcurve.ForNitritedetection,samplesaremixedwiththeColorReagentsAandBandincubatedatroomtemperaturefor5minutes.Thecoloredproductisreadat550–570nm.TheconcentrationofNitriteinthesampleiscalculated,aftermakingasuitablecorrectionforanydilutionofthesample,usingsoftwareavailablewithmostplatereaders.TotalNitricOxidecontentismeasuredafterthesampleisincubatedwithNitrateReductaseandNADH.ThereductaseincombinationwithNADHreducesNitratetoNitrite.Aftera20minuteincubationatroomtemperature,ColorReagentsAandBareaddedandincubatedatroomtemperaturefor5minutes.ThecoloredproductisreadandcalculatedaswiththeNitritedeterminationabove.TheconcentrationofNitrateinthesampleiscalculatedbysubtractingthemeasuredNitriteconcentrationfromtheTotalNitricOxideconcentrationforthesample. | ||||||||||||||||||||||||||||||
KitOverview |
| ||||||||||||||||||||||||||||||
CiteThisProduct | NitricOxideDetectionKit(StressMarqBiosciencesInc.,VictoriaBCCANADA,Catalog#SKT-212) |
BIOLOGicalDescription
AlternativeNames | Nitrogenmonoxide;Nitrogen(II)oxideDetectionKit |
ResearchAreas | CellSignaling,Neuroscience,Post-translationalModifications |
ScientificBackground | Nitricoxide(NO)isadiffusIBLe,transient,reactivemoleculethathasphysiologicaleffectsinthepicomolar-to-micromolarrange.Actingthroughsolubleguanylatecyclaseactivation,NOisanimportantphysiologicalregulatorofthecardiovascular,nervous,andimmunologicalsystems(1).NOisbio-availablebytworoutes.ItcanbeendogenouslygeneratedbyconstitutiveorinducedenzymeslikeNitricOxideSynthaseoritcanbeorallyingestedasnitrates/nitritesforrapiduptakeintocirculationandsubsequentconversion(2).Thereactivenatureofnitricoxideallowsittoactasacytotoxicfactorwhenreleasedduringanimmuneresponsebycellssuchasmacrophages.ThereactivityalsoallowsNOtobeeasilyconvertedtoatoxicrADIcalthatcanproducenitrosativedamagetocells,organellesandmoleculessuchasDNA.Nitrosaylationhowevercanbearegulatedpost-translationalmodificationincellsignaling(3).Thebalanceanddynamicsoftheregulatory/damagefacetsofNOaremajorforcesinmitochondrialsignalinganddysfunction(4).NOislinkednotonlytocoronaryheartdisease,endothelialdysfunctions,erectiledysfunction,andneurologicaldisorders,butalsodiabetes,chronicperiodontitis,autism,cancer,andassortedage-relateddiseases(5-9).ThephysicalpropertiesofNitricOxidemakeitchallengingfordirectdetectionmethods.However,colorimetricmethodscanbeappliedtomeasureitsstablebreak-downproductsnitrate(-NO3)andnitrite(-NO2)(10). |
References | 1.Moncada,SandEAHiggs.(1991)Eur.J.Clin.Invest.,21:361-374. 2.Kapil,V.etal.(2010)Heart,96:1703-1709. 3.Seth,DandStamler,JS.(2007)Curr.Opin.Chem.Biol.,15:1-8. 4.Eursalimsky,JDandMoncada,S.(2007)ATVB27:2524-2531. 5.Knott,ABandBossy-Wetzel,E.(2010)Diab.Obes.Metab.12(Suppl2):126-133. 6.VanDyke,K.etal.(2010)Ann.N.Y.Acad.Sci.1203:138-145. 7.Reher,VSG.etal.(2007)J.OralSci.49(4):271-276. 8.Sogut,S.etal.(2003)Clin.Chim.Acta331:111-117. 9.Balam.E.etal.(2002)Jpn.J.Clin.Oncol.32(5):162-166. 10.Moshage,H.(1997)Clin.Chem.43(4):553-556. |
ProductImages
TypicalStandardCurveforNitricOxideDetectionKitStressXpress®–SKT-212.AssayType:CoupledEnzymeAssay.DetectionMethod:ColorimetricAssay.AssayRange:3.125–200uM.
LinearitywasdeterminedbytakingtwohumanurinesampleswithknownNitriteandTotalNitricOxideConcentrationsandmixingthemingivenratios.Themeasuredconcentrationswerecomparedtotheexpectedvaluesbasedontheratiosused.
ProductCitations(0)
Currentlytherearenocitationsforthisproduct.
ebiomall.com
>
>
>
>
>
>
>
>
>
>
>
>
两个CEX方法A和B测定同一单抗,结果碱性峰比例差不多,酸性峰比例相差约7%,相应主峰也差了7%左右。
具体来说,A方法酸性峰高,主峰低,碱性峰稍微低点;B方法酸性峰低,主峰高,碱性峰稍微高点;另外也做了CIEF,结果呢和A方法更接近。
仔细比较起来,AB两个方法的峰性和数量差不多,就不知道为什么会有这么大的差异。两个方法一个用的WCX柱-磷酸缓冲液,一个用SCX柱-MES缓冲液
大家帮我分析下:
1.两个方法哪个方法更准确,是以酸性峰高的为准还是什么?为什么?
2.这显著差异是由方法造成,具体原因是什么?柱子?
3.CIEF的结果和A方法更接近,是不是可以由此证明A方法更好或者CIEF的方法更好(因为CIEF更快更方便)?
欢迎讨论~
纠正下,A方法用的是Tosoh的柱子,B方法用的是SCX柱。TOSOH的柱子是7um的填料,10cm长。SCX是10um的填料。我本人TOSOH的阳离子柱子用的很少,这次信手用用,结果发现差异很大
那我现在就考虑,在以后方法开发过程中,除了通过流动相pH和组成、梯度、柱子选择来获得样品主峰和酸碱性的最大分离,还要关注各峰比例。因为之前比较方法好坏都只看分离度,尤其是主峰和邻近峰的分离度,获得最大分离度,自然可以做到主峰尽可能纯,但从未认真比较过各峰比例。这是一个大疏忽吧!
另外,CIEF和CEX方法原理还是有点差异的,所以分的是不同的异质体,原液放行两个方法肯定是都要做的。问题就是在早期细胞株筛选和工艺开发阶段,哪个方法才是又快又准。CIEF(iCE280)一般15分钟一个样,比CEX快多了。如果CIEF测得主峰要低于CEX结果,是不是真的完全可以取代CEX呢?CEX分离出的峰远比CIEF的多!
欢迎大家继续讨论~
是否可以理解为纯化水得PH范围为6.3-7.6?能否直接用pH计测量?谢谢!
1.直接用固体磷酸钠配制成50mM的磷酸钠溶液,再调pH到7.4;(我们试着用这个做了下,发现挂不上柱)
2.配置磷酸钠盐缓冲液:按NaH2PO4:Na2HPO4以19:81的摩尔比配制成pH7.4的缓冲液?(附一张百度出来的配方
)
3.如果是磷酸钠盐缓冲液,可以直接将50mM的NaH2PO4的水溶液用NaOH调成pH7.4吗?
再者,2和3这两个方法配制的磷酸钠盐缓冲液有什么区别?最终效果是一样的吗?如果不一样,有什么理论的知识支撑呢?个人感觉是分析化学中酸碱理论中的缓冲液那里的知识。求帮忙解答这些疑问。
另外,我还想问一下,pH对于Ni柱对His-tagged的蛋白的分离纯化影响大吗?是怎么影响的?谢谢大家了!
常用流动相加酸碱后PH的总结,希望大家能够提供一点自己测过的结果,谢谢先
由弱酸及其盐、弱碱及其盐组成的混合溶液,能在一定程度上抵消、减轻外加强酸或强碱对溶液酸碱度的影响,从而保持溶液的pH值相对稳定。这种溶液称为缓冲溶液。
有了源数据之后把源数据按照大小排列,
选中源数据区域-->ALT+A1-->选中图标区右键-->更改图表类型-->散点图
暂无品牌问答