TopoisomeraseIIDrugScreeningKitProductDescription
TopoIIinhibitorshavebeenshowntopossesseffectiveantibacterial,antiviralandanticanceractivity.IdentificationoftopoIIinhibitorsandantagoNISTsrequirestractablebiochemicalassaysemployingpurifiedenzyme. TheTopoisomeraseIIDrugScreeningKitcontainsthereagentsnecessarytoscreenforagentsthataffectenzymaticactivity. Toallowthecustomermaximumflexibility,purifiedtopoisomeraseIImaybepurchasedseparatelyfromthekit(TG2000H). Thiskitwillallowdetectionoftwoclassesoftopoisomeraseinhibitors:thosethatstimulateformationofcleavablecomplexesandthosethatantagonizetopoIIactionontheDNA. Alsoincludedinthiskitisdetaileddocumentationondesignandinterpretationofinhibitiondata. Knowninhibitorsareincludedascontrols. TheDNAsubstrate(pHOT1)includedinthisassayisidealforthesestudiesbecauseitcontainsasingle,highaffinitytopoIIcleavagesite(notethattopoIIisnotparticularlysequencespecific).TheassaysystemisbaseduponevaluatingtheformationofDNAcleavageproductswhichmaybenicked,opencircularDNAorlinearDNA.Theproductsarethenresolvedbyethidiumbromidegelelectrophoresisasdescribedinthekit.
TopoisomeraseIIDrugScreeningKitContents(representativeof100assaykitsize):
-SupercoiledDNA[pHOT1DNA]25ug
-MarkersarelinearpHOT1DNAandopencircularDNA
-10XTopoisomeraseIIassaybuffercomponents*
*ToimprovestABIlity,the10xTopoIIAssaybufferisprovidedastwocomponents: BufferA(noATP)andBufferB(ATP).TheComplete10xAssaybufferismadefresheachtimebycombiningBufferAandB.
-Sodiumdodecylsulfate(10%)
-10XGelloADIngbuffer(bromophenolblue,glycerol)
-Controlinhibitor,Etoposide
-ProteinaseK
-Detailedinstructionmanual
Note:TopoIIenzymeissoldseparately.
TopoIIDrugScreeningBackground:
ThisisacleavagebaseddetectionkitthatwillrevealwhetheraparticularagenteitherantagonizestheactionoftopoIIoraffectsthecleavage/religationcycle. Reactionsareassembledinmicrocentrifugetubesusingreagentssuppliedinthekit(buffer,DNA,controlinhibitor). ThiskitisdesignedtoworkwithpurifiedtopoisomeraseIIprovidedbyTopoGEN. Afteraddingenzyme,thereactionsareincubated15-30min(37°C),andterminatedbyadditionofSDS.
Atypicalreaction:
- H2O:Volumevariable,makeuptovolume(20ulinthiscase)
- 10XAssaybuffer2µl(madefreshfromBuffersAandB,seeKitdetails)
- DNA(supercoiled)1µl
- TestCompoundorcontrolinhibitor:Variable(limitedbysolventeffectsasaddressedincontrols)
- **PurifiedTopoisomeraseII:4-6Units(enzymesoldseparately)
- Incubate15-30minutesat37°C
- Add2µlSDS;digestwithproteinaseK(50ug/ml)
- Add0.1volsampleloadingdye(supplied)
- Loadgel:1%agarosecontaining0.5ug/mlethidiumbromidealongwiththemarkerDNAsincludedinthekit.
ThereactionsindicatedaboveeachlaneinthefigurewerecarriedoutusingtheenzymeprovidedbyTopoGENand1ulofDNA(0.25ugofformIpHOT-1,asprovidedineachkit). Thereactionproductswereanalyzedona1%agarosegel;therunningbuffercontained0.5ug/mlethidiumbromide. Thegelwasthendestainedindistilledwaterpriortophotodocumentation. Formationofthecleavablecomplex(linearDNAspecies)isclearlyvisIBLe. Additionally,relaxationofsupercoiledDNAsubstrate(formIpRYGDNA)wasinhibitedinallreactionsthatcontainedVM26. ThekitcontainsmarkerDNAs,VM-26controlinhibitor,allbuffersandcompleteinstructionsonusingthereagentstoscreenforinhibitorsoftopoisomeraseII. ThemanualspecifiesconditionsforgelelectrophoresistoresolveallDNAforms;publicationqualitygelswillresult.
**CleavagereactionsrequiremoretopoIIthanrelaxationassayssincetheformerisastoichiometricreactionofenzyme. Forexample,theabovereactionscontained4unitsofenzyme(unitsdefinedbydecatenationactivity).
ReviewsandCitations:
- Sudan,S.,&Rupasinghe,H.V.(2014).Flavonoid-EnrichedAppleFractionAF4InducesCellCycleArrest,DNATopoisomeraseIIInhibition,andApoptosisinHumanLiverCancerHepG2Cells.Nutritionandcancer,ahead-of-print(2014):1-10. doi:10.1080/01635581.2014.951733
- ChengMH,YangYC,WongYH,ChenTR,LeeCY,YangCC,ChenSH,YangIN,YangYS,HuangHS,YangCY,HuangMS,ChiuHF:B1,anoveltopoisomeraseIIinhibitor,inducesapoptosisandcellcycleG1arrestinlungadenocarcinomaA549cells. Anti-CancerDrugs2012,23(2):191-199. doi:10.1097/CAD.0b013e32834cd277
- ShiW,MarcusSL,LowaryTL:CytotoxicityandtopoisomeraseI/IIinhibitionofglycosylated2-phenyl-indoles,2-phenyl-benzo[b]thiophenesand2-phenyl-benzy[b]furans. BioorganicandMedicinalChemistry2010. doi:10.1016/j.bmc.2010.10.054
- GongY,FirestoneGL,BjeldanesLF:3,3'-DiindolylmethaneisanoveltopoisomeraseIIacatalyticinhibitorthatinducesS-phaseretardationandmitoticdelayinhumanhepatomaHepG2cells. MolecularPharmacology2006,69(4):1320-1327.
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百奥森18分钟霉菌毒素检测试剂盒，有需要的可以了解下！多种规格型号可供选择
产品名称
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试剂盒在全球市场上的研发与销售呈快速上升趋势,2005年全球市场销售额超过200亿美元,且以15%左右的速度逐年增长.一方面是试剂盒的迅猛发展,而另一方面试剂盒市场良莠不齐的现象愈加明显,试剂盒的生产、销售及认证认可体制尚不完善,没有相应的标准或质量评价政策.且其灵敏度,稳定性及假阴/阳性控制尚不能满足检测需要,采用试剂盒进行检测的公信度受到质疑.
同时,食品安全领域是当前问题最多、最受关注的领域,这个领域的检测包括了物理、化学、微生物及分子生物学基础理论,无论是按检测原理、用途还是其它分类方式,涉及食品安全检测项目的试剂盒的品种是最全最多的.因此,从该领域着手从事评价制度的研究,便于获得基础性数据结果,并由此推广至动植物检疫及其它领域.
随着H7N9禽流感疫情的不断发散,国家流感中心已经多次发放人感染H7N9禽流感检测试剂,覆盖了全国31个流感网络实验室,并表示,诊断试剂的广泛发放是实现关口前移,控制疫情传播、蔓延的重要手段.而一旦H7N9监测关口继续前移,主动监测范围扩大,病毒检测试剂的需求量将进一步加大.
可采用H7N9亚型禽流感病毒RNA检测试剂盒（荧光PCR法）和H7N9禽流感病毒核酸检测试剂盒（PCR-荧光探针法）,定价分别为48人份/盒和48反应/盒,相比市场此前预期的100-200元之间的价格定位低了很多.在检验方法上,卫纪委提醒,前者需要配备全自动荧光PCR检测仪专用PCR扩增管和核酸分离试剂盒(硅胶膜吸附法)等必须设备及咽拭子样本,后者卫计委推荐采用达安基因生产的核酸提取试剂盒进行检验.
我用Promega公司的双荧光素酶检测试剂盒（E2920）检测到的firefly萤光素酶活性很低，只有4*10的3次方；海肾萤光素酶活性有10的6次方。
我用Ad293细胞做了转染，fireflyluciferase质粒：RanillaLuciferase质粒=0.1ug：0.025ug/一个孔（96孔板），共转染了二天，再进行双萤光素酶检测。
我想了解fireflyluciferase活性用promega的这个E2920-双萤光素酶试剂盒检测得到10的3次方，这种数值正常吗？
我做了3次重复实验，每次firefly萤光素酶活性很低，只有4*10的3次方；而海肾萤光素酶活性有10的6次方。
求指教？fireflyluciferase活性低？会是什么原因呢？
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ctl毒性杀伤检测试剂盒是基于LDH在胞浆内含量丰富，正常时不能通过细胞膜，当细胞受损伤或死亡时可释放到细胞外，此时细胞培养液中LDH活性与细胞死亡数目成正比，用比色法测定并与靶细胞对照孔LDH活性比较，可计算效应细胞对靶细胞的杀伤。
牛胰岛素,是一种多肽
在1965年9月17日我国完成了结晶牛胰岛素的全合成。经过严格鉴定，它的结构、生物活力、物理化学性质、结晶形状都和天然的牛胰岛素完全一样。这是世界上第一个人工合成的蛋白质，为人类认识生命、揭开生命奥秘迈出了可喜的一大步。这项成果获1982年中国自然科学一等奖。
1953年，英国人F. SangerSanger由于测定了牛胰岛素的一级结构而获得1958年诺贝尔化学奖。
