
Molecular Weight: | 534.53 |
Formula: | C28H25F3N6O2 |
Purity: | ≥98% |
CAS#: | 1245537-68-1 |
Solubility: | DMSO up to 50 mM |
Chemical Name: | 8-(6-methoxypyridin-3-yl)-3-methyl-1-(4-(3-(trifluoromethyl)piperazin-1-yl)phenyl)-1H-imidazo[4,5-c]quinolin-2(3H)-one |
Storage: | Powder:4oC 1 year. DMSO:4oC3 month;-20oC 1 year. |
Biological Activity:NVP-BGT226 is a novel dual PI3K/mTOR inhibitor with an IC50 ~1 nM. In cellular assays it could produce nearly complete inhibition of PI3K signaling at low nanomolar (<50 nM) concentrations. Flow cytometric analysis revealed an accumulation of cells in the G0–G1 phase with a concomitant loss in the S-phase. TUNEL assay and the analysis of Caspase 3/7 and PARP indicated that BGT226 induced cancer cell death through an apoptosis independent pathway. BGT226 induced autophagy as indicated by the aggregation and upregulation of the microtubule-associated protein light chain 3B-II, and p62 degradation. It is in the phase I/II clinical trials for the treatment of advanced solid tumors.
How to Use:
- In vitro: BGT226 was used at 0.2 µM concentration in cellular assays to investigate its effects on the PI3K/AKT signaling pathways.
- In vivo: BGT226 was dissolved in 90% NMP/10% PEG300 and orally dosed at 5mg/Kg once a day.
Reference:
- 1. Chang KY, et al. Novel phosphoinositide 3-kinase/mTOR dual inhibitor, NVP-BGT226, displays potent growth-inhibitory activity against human head and neck cancer cells in vitro and in vivo. (2011) Clin Cancer Res. 17(22):7116-26.
- 2. Baumann P, et al. Simultaneous targeting of PI3K and mTOR with NVP-BGT226 is highly effective in multiple myeloma. (2012) Anticancer Drugs. 23(1):131-8.
- 3. Sanchez CG, et al. Preclinical modeling of combined phosphatidylinositol-3-kinase inhibition with endocrine therapy for estrogen receptor-positive breast cancer. (2011) Breast Cancer Res. 13(2):R21
- 4. Fokas E, et al. NVP-BEZ235 and NVP-BGT226, dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitors, enhance tumor and endothelial cell radiosensitivity. (2012) Radiat Oncol. 7:48
BGT226_spec.pdf
BGT226_MSDS.pdf
Products are for research use only. Not for human use. |
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该试剂盒是一种时间分辨的荧光共振能量转移免疫分析,该反应是一个竞争免疫反应,即铕标的cAMP示踪复合物与体系中的cAMP竞争结合标有Alexa Fluor 647染料的cAMP抗体。铕标cAMP示踪复合物是通过Biotin标记的cAMP与铕标的抗生物素蛋白链菌素(streptavidin)与抗体的复合物紧密结合产生的。
当抗体结合到示踪剂上时,340nm的激发光激发铕标分子,导致能量转移到Alexa Fluor 647染料上,结果产生665nm的发射光。荧光的强度与样品中的cAMP含量成反比。
本试剂盒用于检测在GPCR激动剂刺激下活细胞或者细胞膜制备品产生的cAMP。对于偶联Gαs的受体,激动剂刺激导致665nm的荧光强度降低,而拮抗剂则可以逆转这一效应;对于偶联Gαi的受体,在激动剂刺激的同时用forskolin刺激cAMP产生,那么激动剂则抑制forskolin诱导的cAMP的生成,因此对照只给forskolin的细胞组可以通过665nm荧光强度的增加反应激动剂的效应。
该试剂盒的灵敏度很高,室温下反应在20h内是稳定的。本试剂盒适用于在384孔板中进行24μl的微量分析。
2.保存条件
避光2~4℃保存,过期时间见外包装。
3.盒内试剂
cAMP标准品:1管,1ml。(50μM)
生物素标记的cAMP(b-cAMP):1管,25μl。
铕标的抗生物素蛋白链菌素:1管,25μl。
荧光标记的cAMP抗体:1管,40μl。
检测缓冲液:1瓶,25ml。
4.需要自配的其他溶液
l Hank’s balanced salt solution (HBSS): NaCl 8.0g、CaCl2 0.14g、KCl 0.4g、 KH2PO4 0.06g、Na2HPO4?7H2O0.09g、MgCl2.6H2O0.10 g、MgSO4.7H2O0.10 g、NaHCO30.35g、葡萄糖1.0g,加H2O至 1000ml (用7.5%NaHCO调节PH值=7.4)
l Versene消化液(1L):EDTA 0.372 g,NaCl 8.0g,KCl 0.20 g,KH2PO40.20g,Na2HPO4 1.15 g,D-glucouse 0.2 g,pH 7.4
l HEPES缓冲液(1mol/L):取2.383gHEPES溶于10ml去离子水中。
l 7.5%BSA溶液:取0.75gBSA溶于10ml去离子水中
l 0.5M IBMX溶液:11.11mg IBMX溶于100μl DMSO中,-20℃冻存。
l 刺激缓冲液(SB):14 ml HBSS(1×)+75μlHEPES(1mol/L)+200μlBSA (7.5%)。(注:在测定细胞cAMP时,反应缓冲液中要加入IBMX 0.5mmol/L)
l 吗啡贮存液(10mM):盐酸吗啡37.585mg溶于10ml生理盐水中,0.22μm滤膜过滤除菌,4℃保存备用。
l 纳络酮母液(100mM):纳络酮4mg溶于100μl 生理盐水中,用时工作液按照1:500稀释,溶剂为含有IBMX的反应缓冲液。
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MaggotsFasterThanScalpelinWoundDebridement
December19,2011—Maggotdebridementtherapy(MDT)appearstobemoreeffectiveforwounddebridementcomparedwithconventionaltherapy,butonlyat1week;afterthattime,anothertypeofdressingshouldbeused,newresearchsuggests.
KristinaOpletalovà,MD,fromtheDepartmentofDermatology,UniversityofCaen,France,andcolleaguespublishedonlineDecember19intheArchivesofDermatology.
MedicalmaggotswereapprovedbytheUSFoodandDrugAdmiNISTrationasamedicaldeviceforwounddebridementin2004.Accordingtotheresearchers,useofmaggotsintreatingwoundsisassociatedwitheffectivewounddebridement,antibacterialeffects,andstimulationofwoundhealing.
However,theypointout,"[r]elativelyfewclinicalstudieshavebeenconductedandtheresultsarenotclear,partlyowingtomethodologicassessmentproblems."
InthecurrentProspective,randomizedcontrolled,phase3clinicaltrial,theresearcherssoughttodeterminetheefficacyofbaggedlarvaeonwounddebridementincomparisonwithconventionaltreatment.
TheprimaryobjectivewastocomparethemeanpercentageofsloughinwoundstreatedwithMDTwiththatofconventionaltreatmentatday15.Thestudyincluded119patientswithanonhealing,sloughywoundthatwas40cm2orsmallerandlessthan2cmdeep.Patientsalsohadananklebrachialindexof0.8orhigher.
Treatmentwasadministeredduringa2-weekhospitalstay.Conventionaltreatmentconsistedofsurgicaldebridement3timesaweekwithascalpel,withuseoftopicalanesthesia.TheMDTwasadministeredusinganencloseddressing(Vitapad,BioMondeLaboratories)containing80sterilemaggots.Atdischarge,aconventionaldressingwasapplied,andpatientswerefollowed-upatday30.
DebridementbyMDTwassignificantlyfasterthansurgicaldebridementduringthefirstweekoftreatment,reachingthesamelevelthecontrolgroupreachedatday15.NobenefitforMDTcomparedwithconventionaltreatmentinhealingrateswasobserved.Atday8,54.5%intheMDTgroupvs66.5%inthecontrolgroup(P=.04)hadevidenceofsloughandwoundhealing.However,byday15,themeanpercentageofsloughwas55.4%intheMDTgroupand53.8%inthecontrolgroup(P=.78).
"AthoughMDTshowsnosignificantbenefitatday15comparedwithconventionaltreatment,debridementbyMDTissignificantlyfasterandoccursduringthefirstweekoftreatment,"theresearchersconclude."Becausethereisnobenefitincontinuingthetreatmentafter1week,anothertypeofdressingshouldbeusedafter2or3applicationsofMDT."
Painscoresweresimilarandmildinbothgroups,althoughincontrasttoconventionaltreatment,MDTwasperformedwithouttopicalanesthesia.
Accordingtotheresearchers,noneofthepatientswerereticentaboutundergoingMDT."[A]crawlingsensationonthewoundwasrarelyandalmostequallynotedinbothgroups,revealingthatthesensationwassubjective,"Dr.Opletalovàandcolleaguespointout.
TwoquestionsregardingMDTremainunanswered,theauthorsnote."Candebridementbeimprovedusingmoremaggotsperdressing?Ifso,wouldthesedressingsbemorepainful?Furtherstudiesareneededtoanswerthesequestions."
ThestudywassupportedbygrantsfromtheClinicalResearchHospitalProgramandfromtheFrenchSocietyofDermatology.Theauthorshavedisclosednorelevantfinancialrelationships.
该试剂盒是一种时间分辨的荧光共振能量转移免疫分析,该反应是一个竞争免疫反应,即铕标的cAMP示踪复合物与体系中的cAMP竞争结合标有Alexa Fluor 647染料的cAMP抗体。铕标cAMP示踪复合物是通过Biotin标记的cAMP与铕标的抗生物素蛋白链菌素(streptavidin)与抗体的复合物紧密结合产生的。
当抗体结合到示踪剂上时,340nm的激发光激发铕标分子,导致能量转移到Alexa Fluor 647染料上,结果产生665nm的发射光。荧光的强度与样品中的cAMP含量成反比。
本试剂盒用于检测在GPCR激动剂刺激下活细胞或者细胞膜制备品产生的cAMP。对于偶联Gαs的受体,激动剂刺激导致665nm的荧光强度降低,而拮抗剂则可以逆转这一效应;对于偶联Gαi的受体,在激动剂刺激的同时用forskolin刺激cAMP产生,那么激动剂则抑制forskolin诱导的cAMP的生成,因此对照只给forskolin的细胞组可以通过665nm荧光强度的增加反应激动剂的效应。
该试剂盒的灵敏度很高,室温下反应在20h内是稳定的。本试剂盒适用于在384孔板中进行24μl的微量分析。
抑制剂刺激细胞后,需要用PBS清洗后再做后续实验吗
、
Cell-Based ELISA
的优点:
Cell-Based ELISA
(基于细胞的
ELISA
)是一种全新的
ELISA
技术,有两个最突出的优点:
1.1
不需要抽提蛋白、包被微孔板:细胞直接在微孔板里培养,待检测的时候,将细胞固定在微孔板上并
进行通透处理即可。这样就避免了抽提蛋白时,由于客观和主观上引起样品的损失而导致实验结果在一定
程度上偏离了实际情况。
同时不用包被微孔板,
简化了实验流程,
有助于提高效率。
科研人员在一个
96
孔
酶联板上,便能检测目标细胞蛋白经刺激或抑制作用后的表现。由于省去抽提蛋白和裂解细胞的步骤,样
本的损失也能降到最低,比起其他普通的
ELISA
测定方法,这项全新的
ELISA
技术能更快速、更方便
地一次检测大量的细胞内蛋白。
1.2
可同时检测两种不同蛋白:
封闭后加入两种抗不同蛋白且来源于不同宿主的一抗,
然后再加入不同的
二抗,加入两种荧光底物,检测两个波长。同时检测两种蛋白的好处是显而易见的,可以减少工作量,此
外还可以满足一些特殊的实验需要,例如,需要测定某个蛋白的磷酸化比例,就需要测定磷酸化蛋白的数
量和总蛋白的数量,这两个测定在同一次实验进行,有助于消除实验误差,得到更为精确的实验结果。
2
、
Cell-Based ELISA
的两种技术:
某些公司发展了双通道
Cell-Based ELISA
技术;
双通道与单通道
Cell-based ELISA
比较:
双通道
cell-based ELISA
,顾名思义,即一次可以同时检测两种目的蛋白,
R&D Systems
提供的
Cell-based ELISA
就是双通道
cell-based ELISA
,原理略:(需要荧光检测方式和相应仪器);
单通道
cell-based ELISA
,即一次只能检测一种目的蛋白,他和普通
ELISA
的主要区别在于样品处理过
程
3
、
cell-based ELISA
的应用:
该产品,最多发展起来的是用于检测磷酸化和非磷酸化蛋白的相对含量;
4
、有
cell-based ELISA
产品的公司:目前,多家
elisa
产品提供
商均
提供
cell
based
elisa
试剂盒,
如
Rnd
systems
,
raybiotech
,
ebioscience
,
millipore
等公司;
5
、如何自己进行
cell based elisa
实验?
事实上,根据单通道的
cell based elisa
原理,可以自己建立
cell based elisa
实验系统;
细胞加入培养板中;
加入刺激物或者抑制剂进行培养;
对细胞进行固定或者封闭;
加入第一抗体;
加入
HRP
结合的二抗(二抗的选择,同
western blot
);
加入底物进行显色;

