| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 405/None |
| MW | N/A |
| CAS# | N/A |
| Solvent | N/A |
| Storage | F/D/L |
| Category | EnzymeDetection Phosphatases |
| Related | BiochemicalAssays |
1.Prepare100XpNPPstocksolutions:
Add300µLofdistilledH2OintothevialofpNPP(ComponentA).Mixthereagentswell.ThepNPPstocksolutionshouldbeusedpromptly.Anyremainingsolutionshouldbealiquotedandrefrozenat-20oC.
Note:Avoidrepeatedfreeze-thawcycles.Thesolutionshouldbegoodfor3-4weeksifstoredat-20oC.
2.PreparepNPPreactionmixture:
PreparepNPPreactionmixtureaccordingtothefollowingtableandkeepfromlight.
Table1.pNPPReactionMixtureforone96-wellplate
Components | Volume |
pNPPstocksolution(100X,fromStep1) | 50µL |
AssayBuffer(ComponentB) | 5mL |
Totalvolume | 5mL |
Note:Preparefreshreactionmixtureforeachexperiment.
3.Prepareseriallydilutedalkalinephosphatasestandards(0to100mU/mL):
3.1 Add100μLofdistilledH2Owith0.1%BSA(H2O-0.1%BSA)toAlkalinePhosphataseStandard(ComponentC,10units)togeneratea100units/mLAlkalinePhosphatasestandardsolution.
Note:Thealkalinephosphatasestandardsolutionisnotstable.Theunusedsolutionshouldbealiquotedandstoredat-20oC.Avoidrepeatedfreeze-thawcycles.
3.2 Add10μLof100units/mLAlkalinePhosphataseStandardsolution(fromStep3.1)to990µLofH2O-0.1%BSAtogeneratea1,000mU/mLAlkalinePhosphataseStandardsolution.
3.3 Take100μLof1,000mU/mLAlkalinePhosphataseStandardsolution(fromStep3.2)toperform1:10andthen1:3serialdilutionstoget100,30,10,3,1,0.3,0.1,and0mU/mLseriallydilutedAlkalinePhosphataseStandards.
3.4 Addseriallydilutedalkalinephosphatasestandardsandalkalinephosphatasecontainingtestsamplesintoawhite/clearbottom96-wellmicroplateasdescribedinTables2and3.
Note1:Preparecellsortissuesamplesasdesired.
Note2:Unusedportionofdilutedalkalinephosphatasestandardsolutionshouldbediscarded.
Table2LayoutofAlkalinephosphatasestandardsandsamplesinawhite/clearbottom96-wellmicroplate
BL | BL | TS | TS | …. | …. |
|
|
|
|
|
|
AS1 | AS1 | …. | …. | …. | …. |
|
|
|
|
|
|
AS2 | AS2 |
|
|
|
|
|
|
|
|
|
|
AS3 | AS3 |
|
|
|
|
|
|
|
|
|
|
AS4 | AS4 |
|
|
|
|
|
|
|
|
|
|
AS5 | AS5 |
|
|
|
|
|
|
|
|
|
|
AS6 | AS6 |
|
|
|
|
|
|
|
|
|
|
AS7 | AS7 |
|
|
|
|
|
|
|
|
|
|
Note:AS=AlkalinePhosphataseStandards;BL=BlankControl;TS=TestSamples
Table3Reagentcompositionforeachwell
AlkalinePhosphataseStandards | BlankControl | TestSample |
SerialDilutions*:50μL | H2O-0.1%BSA:50μL | 50μL |
Note:Addtheseriallydilutedalkalinephosphatasestandardsfrom100to0.01mU/mLintowellsfromAS1toAS7induplicate.
4.Runalkalinephosphataseassayinsupernatants:
4.1 Add50μLofassayreactionmixture(fromStep2)intoeachwellofalkalinephosphatasestandard,blankcontrol,andtestsamples(seeStep3.4,Table3)tomakethetotalalkalinephosphataseassayvolumeof100µL/well.
Note:Fora384-wellplate,add25μLofsampleand25μLofassayreactionmixtureintoeachwell.
4.2 Incubatethereactionatthedesiredtemperaturefor10to30minutes,protectedfromlight.
4.3 Monitortheabsorbanceincreasewithanabsorbanceplatereaderat400nm.
5.Runalkalinephosphataseassayincells:
5.1 Treatthecellsasdesired.
5.2 Addequalvolumeofassayreactionmixture(fromStep2)intoeachcellwell(suchas100μL/96-wellplate,or50μL/384-wellplate).
Note:Alternatively,removethegrowthmediumfromthecellplate,andmake1:1dilutionofthe5mLassayreactionmixture(fromStep2,Table1)with5mLdistilledH2O.ThenAdd100μL(fora96-wellplate)or50uL(fora384-wellplate)of1:1dilutedassayreactionmixturetothecellwells(fromStep5.2).
5.3 Incubatethereactionatthedesiredtemperaturefor30to60minutes,protectedfromlight.
5.4 Monitortheabsorbanceincreasewithanabsorbanceplatereaderat400nm.
| References&Citations |  CitationExplorer |
Theimpactofvariousscaffoldcomponentsonvascularizedboneconstructs
Authors:AhmadEweida,MatthiasSchulte,OliverFrisch,UlrichKneser,LeilaHarhaus
Journal:JournalofCranio-MaxillofacialSurgery(2017)
AMineralizedHighStrengthandToughHydrogelforSkullBoneRegeneration
Authors:BingXu,PengbinZheng,FeiGao,WeiWang,HongtaoZhang,XuranZhang,XuequanFeng,WenguangLiu
Journal:AdvancedFunctionalMaterials(2016)
DRGaxonelongationandgrowthconecollapserateinducedbySema3AaredifferentlydependentonNGFconcentration
Authors:AndriusKaselis,RimantasTreinys,RutaVosyliute,SauliusSatkauskas
Journal:Cellularandmolecularneurobiology(2014):289--296
Acuteoraltoxicityandkineticbehaviorsofinorganiclayerednanoparticles
Authors:JinYu,Hea-EunChung,Soo-JinChoi
Journal:JournalofNanomaterials(2013):12
EffectofSomeAntihypertensiveDrugsonAlkalinePhosphataseandDNAofMice
Authors:OYEl-Khawaga,AEl-Waseef,YOEllazec,MMEl-Naggar,ABDMAlla
Journal:InternationalJournalofGenomicsandProteomics(2013):60
Anengineeringunderstandingofthesmallintestine
Authors:MonicaRosaliaJaimeFonseca
Journal:(2012)
Cytotoxicityandalkalinephosphataseactivityevaluationofendosequencerootrepairmaterial
Authors:MahmoudRezaModareszadeh,PeterMDiFiore,DavidATipton,NargesSalamat
Journal:Journalofendodontics(2012):1101--1105
Alginate-loadedliposomescanprotectencapsulatedalkalinephosphatasefunctionalitywhenexposedtogastricpH
Authors:AlanMSmith,MonicaRJaime-Fonseca,LiamMGrover,SerafimBakalis
Journal:Journalofagriculturalandfoodchemistry(2010):4719--4724
Regulationoftheosteoblast-specifictranscriptionfactorOsterixbyNO66,aJumonjifamilyhistonedemethylase
Authors:KrishnaMSinha,HideyoYasuda,MadeleneMCoombes,SharonYRDent,BenoitDeCrombrugghe
Journal:TheEMBOjournal(2010):68--79
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 |
本网站将在规定时间内给予删除等相关处理。
