Overview | ![]() PrinterFriendlyVersion |
Ex/Em(nm) | 571/585 |
MW | N/A |
CAS# | N/A |
Solvent | N/A |
Storage | F/D/L |
Category | EnzymeDetection Transferases |
Related | BiochemicalAssays |
1.PrepareserialdilutionsofASTstandard(1to300mU/mL):
1.1 Add100µLDPBSBuffertoASTPositiveControl(ComponentD)tomake100U/mLASTstandardsolution.
Note:TheunusedASTPositiveControl(ComponentD)shouldbedividedintosinglueusealiquotsandstoredat-20oC.
1.2 Add3μLof100U/mLASTstandardsolution(fromStep1.1)into997µLDPBSbufferwith0.1%BSAtogenerate300mU/mLASTstandardsolution.
1.3 Take300μLof300mU/mLASTstandardsolutiontoperform1:3serialdilutionstoget100,30,10,3,1,0.3and0mU/mLserialdilutionsofASTstandard.
1.4 AddserialdilutionsofASTstandardandASTcontainingtestsamplesintoasolidblack96-wellmicroplateasdescribedinTables1and2.
Note:DilutethetestsamplestotheconcentrationrangeinDPBSbufferwith0.1%BSAifneeded.
Table1LayoutofASTstandardsandtestsamplesinasolidblack96-wellmicroplate
BL | BL | TS | TS | …. | …. |
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AST1 | AST1 | …. | …. | …. | …. |
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AST2 | AST2 |
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AST3 | AST3 |
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AST4 | AST4 |
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AST5 | AST5 |
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AST6 | AST6 |
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AST7 | AST7 |
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Note:AST=ASTStandards,BL=BlankControl,TS=TestSamples.
Table2Reagentcompositionforeachwell
ASTStandard | BlankControl | TestSample |
SerialDilutions*:50μL | DPBSwith0.1%BSA:50μL | 50μL |
Note1:AddtheseriallydilutedASTstandardsfrom0.3mU/mLto300mU/mLintowellsfromAST1toAST7induplicate.
Note2:TheASTstandardsareforpositivecontrolonly,andshouldnotbereliedonasaquantitationstandardforenzymeactivity.
2.PrepareASTassaymixture:
2.1Add100μLofddH2OintothevialofNAD(ComponentC)tohave100XNADsolution.
2.2Add10mLofASTAssayBuffer(ComponentB)intothebottleofASTEnzymeMixture
(ComponentA),andmixwell.
2.3Addwholevialof100XNADsolution(fromStep2.1)intotheASTEnzymeMixturesolution(fromStep
2.2)tohaveASTassaymixture.
Note1:ThisASTassaymixtureisenoughfortwo96-wellplates.Itisunstableatroomtemperature,andshouldbeusedpromptlywithin2hoursandavoidexposuretolight.
Note2:Alternatively,onecanmakea50XofASTEnzymeMixturestocksolutionbyadding200μLofH2OintothebottleofComponentA,andthenpreparetheASTassaymixturebymixthestocksolutionwithassaybuffer(ComponentB)and100xNADsolutionproportionally. Aliquotandstoretheunused50XASTEnzymeMixturestocksolutionand100XNADsolutionat-20oC,andavoidfreeze-thawcycles.
3.RunASTassay:
3.1 Add50μLofASTassaymixture(fromStep2.3)toeachwellofASTstandard,blankcontrol,andtestsamples(seeStep1.4)tomakethetotalASTassayvolumeof100µL/well.
Note:Fora384-wellplate,add25μLofsampleand25μLofASTassaymixtureintoeachwell.
3.2 Incubatethereactionat37°Cfor20-30minutesorroomtemperaturefor60minutes,protectedfromlight.
Note:ThebackgroundofBlankControlincreaseswithtimeandtemperature.
3.3 MonitorthefluorescenceincreasewithafluorescenceplatereaderatEx/Em=530-570/590-600nm(optimalEx/Em=540/590nm,cutoffat570nm).
References&Citations | ![]() CitationExplorer |
Invitroandinvivostudyoftheapplicationofvolvoxspherestoco-culturevehiclesinlivertissueengineering
Authors:SiouHanChang,HanHsiangHuang,PeiLeunKang,YuChianWu,Ming-HuangChang,ShyhMingKuo
Journal:ActaBiomaterialia(2017)
AAT Bioquest常用产品报价单
产品名 | 货号 | 公司分类 | 商城分类 | 美金价 |
AAT Bioquest/Fluorescein, disodium salt *CAS 518-47-8*/2/100 mg | 2 | iFluor Rapid Tests | 酸碱缓冲液 | 1088.00 | AAT Bioquest/14-3-3 ζ (Ab-58) Antibody/8B0001/50 ug | 8B0001 | Signaling Intermediate Ab | 重组抗体 | 2828.00 | AAT Bioquest/Opioid Receptor (Phospho-Ser375) Antibody/8A0022/50 ug | 8A0022 | Phospho-Specific Ab | 受体 | 2828.00 | AAT Bioquest/iFluor™ 350 goat anti-mouse IgG (H+L)/16440/200 ug | 16440 | Fluorescent Anti-IgGs | 荧光染料 | 1088.00 | AAT Bioquest/trFluor™ Eu goat anti-mouse IgG (H+L)/16518/100 ug | 16518 | Fluorescent Anti-IgGs | 抗小鼠 | 4278.00 | AAT Bioquest/RPE-streptavidin conjugate/16900/100 ug | 16900 | Streptavidin Conjugates | 其他生物染料 | 1378.00 | AAT Bioquest/GCNT3 Antibody/8C14708/50 ug | 8C14708 | Signaling Intermediate Ab | 功能性抗体 | 2828.00 | AAT Bioquest/Amplite™ Luciferase Reporter Gene Assay Kit *Bright Glow*/12518/1 plate | 12518 | Reporter Gene Enzymes | 其它检测试剂盒 | 2103.00 | AAT Bioquest/ReadiLink™ Rapid mFluor™ Violet 450 Antibody Labeling Kit *Microscale Optimized for Labeling 50 &mi | 1100 | mFluor™ Dyes and Kits | 标记试剂盒 | 2103.00 | AAT Bioquest/Amplite™ Colorimetric Urea Quantitation Kit *Blue Color*/10058/200 Tests | 10058 | Diagnostic Molecules | 定量试剂盒 | 2828.00 | AAT Bioquest/Screen Quest™ CHO-Gqi Chimera Cell line/38101/Each | 38101 | cAMP GPCR Assays | 常规细胞株 | 0.00 |