Description:
TheTechnozym®tPACombiActibind®ELISAtestkitmeasurest-PAfunctionalactivity,non-complexedt-PAantigenandt-PA-PAI-1complexes.Itisnotaffectedbythepresenceofotherplasminogenactivators.Themeasurementrangeis0.05-10IU/mLfort-PAactivityand0.1-20ng/mLfort-PAantigen.Theinter-andintra-assayvariationsarelessthan10%and5%,respectively.
KitComposition:
Determination:42samplesinduplicate
- PLATE+PLATECOVER12x8wellplasticmicrotitrestripsprecoatedwithamonoclonalanti-t-PAcoatingantibodyinbicarbonatebuffer,1%bovineserumalbumin(BSA),(TC-CodeGN).
- STANDARD1xlyophilizedrecombinantt-PA,calibratedagainstNIBSC86/670(TC-CodeAW).
- POX-ANTIBODY1xconjugatedantit-PAantibody(concentrated),0,3ml.(TC-CodeKM).
- INCUBATIONBUFFER1×90ml(PBS;pH7,3)containsstABIliserprotein,0,05%proclinandbluedye,1bottle,,readyforuse(TC-CodeNB)
- SUBSTRATE–(greencap)1x12mLTMB(tetramethylbenzidine)(TC-CodeKN)readytouse.
- STOPSOLUTION–(redcap)1x15mLsulphuricacid0.45mol/l(TC-CodeKK)readytouse.
- WASHINGBUFFER–concentrate(1+11,5)1x80ml(PBSpH7,3)containingdetergent,0,01%merthiolat(TC-CodeNA).
- PLASMINOGENACTIVATORDETECTIONMIXTURE.1xlyophilizedH-D-norleucyl-hexahydrotyrosyl-lysine-p-nitroanilidediacetatesalt,GluplasminogenandCyanogenbromidefibrinogenfragments(TCCodeBA).
- PLASMIN-ACTIVATORDETECTIONMIXTUREDILUTIONBUFFER(whitecap)1x20mL50mMTris,12mMNaCl.Readytouse(TC-CodeDA).
Alsorequired
- MicroPipettesandamultichannelmicropipette;pipettetips.
- Glassorplastictesttubesfordilutingthestandard+samples.
- Laboratorybottlesorbeakersandgraduatedcylindersfordilutingwashandincubationbuffer.
- Distilledordeionisedwater.
- Absorbentpapertowels.
- Microtitreplatewasher(alternatively,washingcanbeperformedmanuallyusingamultichannelpipetteorrepeatingsyringe).
- Amicrotitrereaderequippedwitha405nmfilterand,ifpossIBLe,a492nmreferencefilter.
- A37°Cincubator
- Graphpaper.
MeasurementPriniple:
IntheActibindt-PAtest,anantibody,whichdoesnotinterferewitht-PAfunctionalactivity,iscoatedontoamicrotitreplateandusedtobindt-PAcontainedinplasmatotheplatesurface.
Followinganincubationperiod,non-boundplasmacomponentsarewashedawayandanactivitysubstratesolutioncontainingGlu-plasminogen,CNBr-fragmentsoffibrinogenandachromogenicplasminsubstrateisincubatedintheplate.Theboundt-PAactivatesGlu-plasminogentoyieldplasmin.Thereactionbetweenplasminandthechromogenicplasminsubstratereleasesacolouredproductwhoseconcentrationisproportionaltotheamountofactivet-PAinthetestsample.
Afterphotometricallymeasuringthisreaction,themicrotitreplateiswashedtoremovetheactivitysubstratesolution.Thet-PAantigenremainsboundtotheplate.AhorserADIshperoxidase(POX)conjugatedmonoclonalanti-t-PAantibodywhichrecognizesbothactive,t-PAandinactivet-PAisthenincubatedontheplate.Followingincubationandwashingoftheplate,aPOXsubstrateisusedtoproduceacolouredreactionproductwhoseconcentrationisproportionaltothetotalt-PAcontentofthetestsample.
Background:
Tissueplasminogenactivator(t-PA)isa70,000daltonglycoproteinwhichservesasthemajoractivatorofthebloodfibrinolyticsystem.Itissynthesizedprincipallybyendothelialcellsandreleasedintothebloodstreamfollowingstimulus,e.g.exerciseorvenousocclusion.t-PAisuniqueamongtheplasminogenactivatorsduetothefactthatitsactionishighlyfibrinspecific.Onlyinthepresenceoffibrin,towhichitbinds,doest-PAefficientlyactivateGlu-plasminogentoyieldthecleavageproduct,plasmin.Plasmin,inturn,degradesfibrin,theinsolubleproteinpolymerinthrombi.
tPAcirculatesinplasmaataconcentrationofapproximately2-8ng/mL.However,95%ofcirculatingt-PAiscomplexedwiththeplasminogenactivatorinhibitor,PAI-1,andthusinaninactiveform.Uponvenousocclusiontheconcentrationoft-PAantigenincreasesto15ng/mLorhigherandt-PA’sactivityrisesaccordingly.Thisincreaseisprobablyduetoacombinationofincreasedt-PAreleasebyendothelialcellsandareductionint-PAclearancefromtheocclusionsite.t-PAactivitycanonlybemeasuredinsamplesfromwhichinhibitorsoft-PAhavebeenremoved.IntheActibindsystemthisissimplyaccomplishedbyspecificimmunoabsorptionofthetPAcontainedintheplasmasamplebyanantibodyimmobilizedontoamicrotitreplatesurface.Inthisfirststept-PAandt-PA-PAI-1-complexesareboundtotheplateandactive,non-complexedt-PAistherafterquantifiedusingasolutionofGlu-plasminogen,fibrinogenfragmentsandasyntheticsubstratewhichreactswithplasminformedinthereactiontoyieldacolouredreactionproduct.Lowactivityinthisfirststepcanbeduetoeitheradecreaseinthequantityoftotalt-PAboundtotheplateortoanexcessofPAI-1intheplasmasample,whichinactivatest-PAandleadstotheformationoft-PA-PAI-1-complexes.Todifferentiatebetweenthesetworeasonsfordecreasedt-PAfunction,thesecondpartoftheActibindassayisthenusedtodeterminethetotalamountoft-PAantigenboundontothemicrotiterplatethroughtheuseofaperoxidase-labelledmonoclonalantibody.
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2.NaHCO3,Ba(OH)3,H2SO4
3.HCL,NaAlO2,NaHSO4
4.Ca(OH)2,Na2CO3,BaCO3
谢谢了
要原因
请问有什么方法检测细胞内活性氧更好，各位有什么经验，及相关试剂盒推荐一下。
A.H2SO4Na2SO4AgNO3BaCl2.
B.NaOHNa2CO3NaHSO4MgCl2
C.CaCl2NaNO3MgSO4BaCl2
D.HNO3KOHKClK2CO3
请问下有无同学需要H37RA的？我是做EAE模型的，上个月购买了BDDifco公司的H37RA（货号），因为购买的时候只能整盒6支购买，但我们用不了那么多，所以想问问有无同学需要的，100mg/支，800元/支或用等价试剂交换。地址广州。有需要的请私信，谢谢！
（1）优级纯试剂 亦称保证试剂，为一级品，纯度高，杂质极少，主要用于精密分析和科学研究，常以GR表示。
（2）分析纯试剂 亦称分析试剂，为二级品，纯度略低于优级纯，杂质含量略高于优级纯，适用于重要分析和一般性研究工作，常以AR表示。
（3）化学纯试剂 为三级品，纯度较分析纯差，但高于实验试剂，适用于工厂、学校一般性的分析工作，常以CP表示。
（4）实验试剂 为四级品，纯度比化学纯差，但比工业品纯度高，主要用于一般化学实验，不能用于分析工作，常以 LR表示。
以上按试剂纯度的分类法已在我国通用。根据化学工业部颁布的“化学试剂包装及标志”的规定，化学试剂的不同等级分别用各种不同的颜色来标志，见表1。
表1 我国化学试剂的等级及标志
bhclna2so4nano3na2co3
chclnaohna2co3nacl
dba(oh)2nahco3alcl3nahso4
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