| Overview | PrinterFriendlyVersion |
| Ex/Em(nm) | 571/585 |
| MW | N/A |
| CAS# | N/A |
| Solvent | N/A |
| Storage | F/D/L |
| Category | SmallMoleculeDetection DiagnosticMolecules |
| Related | BiochemicalAssays |
| Spectrum | AdvancedSpectrumViewer |
1.Preparestocksolutions:
1.1 Amplite™Redstocksolution(250X):Add40µLofDMSO(ComponentE)intothevialofAmplite™Red(ComponentA)tomakea250Xstocksolution.
Note:TheunusedAmplite™Redstocksolutionshouldbedividedintosingleusealiquots.Storeat-20oCandavoidexposuretolight.
1.2 Cholinestocksolution:Add400µLofddH2OintothevialofCholineStandard(ComponentC)tomake50mMcholinestocksolution.
Note:Theunusedcholinestocksolutionshouldbedividedintosingleusealiquotsandstoredat‑20 oC.
2.Preparecholineassaymixture:
2.1 Add5mLofAssayBuffer(ComponentD)tothebottleofCholineProbe(ComponentB)andmixwell.
2.2 Add20µLofAmplite™Redstocksolution(250X,fromStep1.1)intotheCholineProbebottle(fromStep2.1)tomakethecholineassaymixture.
Note:Thecholineassaymixtureshouldbeusedpromptlyandkeptfromlight.Theassaybackgroundwouldincreasewithlongerstoragetime.
3.Prepareserialdilutionsofcholinestandard(0to30μM):
3.1 Add20μLof50mMcholinestandardstocksolution(fromStep1.2)to980µLAssayBuffer(ComponentD)togenerate1000μMstandardsolution.
Note:Dilutedcholinestandardsolutionisunstable,andshouldbeusedwithin4hours.
3.2 Take30μLof1000μMstandard(fromStep3.1)to970µLAssayBuffer(ComponentD)togenerate30μMcholinestandardsolution,andthenperform1:3serialdilutionstoget10,3,1,0.3,0.1,0.03,and0μMcholinestandard.
3.3 Addcholinestandardsandcholinecontainingtestsamplesintoa96-wellsolidblackmicroplateasdescribedinTables1and2.
Note:Treatthecellsortissuesamplesasdesired.
Table1.Layoutofcholinestandardsandtestsamplesinasolidblack96-wellmicroplate*
BL | BL | TS | TS | …. | …. |
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CS1 | CS1 | …. | …. | …. | …. |
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CS2 | CS2 |
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CS3 | CS3 |
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CS4 | CS4 |
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CS5 | CS5 |
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CS6 | CS6 |
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CS7 | CS7 |
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*Note:CS=CholineStandards;BL=BlankControl;TS=TestSamples
Table2.Reagentcompositionforeachwell*
CholineStandard | BlankControl | TestSample |
Serialdilutions*(50μL) | Assaybuffer:50μL | 50μL |
*Note:Addtheseriallydilutionsofcholinestandardsfrom0.03to30μMintowellsfromCS1toCS7induplicate.
4.Runcholineassay:
4.1 Add50μLofcholineassaymixture(fromStep2.2)toeachwellofthecholinestandard,blankcontrol,andtestsamples(seeStep3.3)tomakethetotalcholineassayvolumeof100µL/well.
Note:Fora384-wellplate,add25μLsampleand25μLofcholineassaymixtureperwell.
4.2 Incubatethereactionatroomtemperaturefor10to30minutes,protectedfromlight.
4.3 MonitorthefluorescenceincreasewithafluorescencemicroplatereaderatEx/Em=540/590nm(cutoffat570nm).
| References&Citations | CitationExplorer |
StructuralsnapshotsofthecatalyticcycleofthephosphodiesteraseAutotaxin
Authors:JensHausmann,Willem-JanKeune,AgnesLHipgraveEderveen,LeonievanZeijl,RobbiePJoosten,AnastassisPerrakis
Journal:JournalofstructuralBIOLOGy(2016)
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 |



