- BMS-708163 (Avagacestat)
- DAPT (GSI-IX)
- BMS 299897
- YO-01027 (Dibenzazepine, DBZ)
- Flurizan
- Semagacestat (LY450139)
LY-411575Gamma secretase inhibitor |
Sample solution is provided at 25 µL, 10mM.
Quality Control & MSDS
- View current batch:
- Purity = 99.10%
- COA (Certificate Of Analysis)
- HPLC
- NMR (Nuclear Magnetic Resonance)
- MSDS (Material Safety Data Sheet)
- Datasheet
Chemical structure
Description | LY411575 is a potent γ-secretase inhibitor with IC50 value of 0.078 nM and 0.082 nM for membrane and cell-based, respectively. | |||||
Targets | γ-secretase | Notch S3 cleavage | ||||
IC50 | 0.078 nM (membrane-based) 0.082 nM (cell-based) | 0.39 nM |
Kinase experiment [1]: | |
Inhibitory activities | Procedures for measuring γ-secretase activity in membranes prepared from HEK293 cells expressing APP. Intact HEK293 cells expressing either APP or NE were treated with various concentrations of LY-411,575 for 4 h at 37℃. In the case of cells expressing NE, cells were lysed, the cell lysates were separated on a 4-12% NuPAGE gel, and the processed NICD fragment was detected via Western blot with a cleavage site-specific antibody. The inhibition of NICD production was quantified by spot densitometric analysis using FluorChem. In the case of cells expressing APP, the conditioned medium was collected, centrifuged at 10,000×g for 5 min to remove cell debris, and stored at -20℃ prior to the determination ofAβ levels. Aβ40 and -42 produced in HEK293 membrane- and cell-based assays, as well as plasma Aβ40 and cortex Aβ40 from TgCRND8 mice, were analyzed without pretreatment using an electrochemiluminescence detection-based immunoassay. Plasma Aβ42 was measured by enzyme-linked immunosorbent assay. A commercially available enzyme-linked immunosorbent assay kit was used to measure cortex Aβ42. |
Cell experiment [1]: | |
Cell lines | HEK293 cells expressing human APP carrying both the Swedish and London mutations or NE. |
Preparation method | Soluble in DMSO > 10 mM. General tips for obtaining a higher concentration: Please warm the tube at 37℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months. |
Reaction Conditions | 4 h. |
Applications | In HEK293 cells expressing human APP or N E, LY-411,575 inhibits Aβ40 and NICD production with IC50 values of 0.082 and 0.39 nM, respectively. |
Animal experiment [1]: | |
Animal models | Six-week-old female TgCRND8 or male C57BL/6 mice. |
Dosage form | 1-10 mg/kg; dosed orally once/day for 5 or 15 days. |
Preparation method | Formulated as 10 mg/ml solutions in 50% polyethylene glycol, 30% propylene glycol, 10% ethanol and diluted in 0.4% methylcellulose for dosing. |
Applications | In TgCRND8 mice, LY-411,575 decreases brain and plasma Aβ40 and Aβ42. LY-411,575 (10 mg/kg) reduces weight of mice by 2 g. LY-411,575 induces a marked atrophy of the cortical zone of the thymus and reduces the amount of thymocyte cells. |
Other notes | Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal. |
References: [1]. Wong GT, Manfra D, Poulet FM, et al. Chronic treatment with the gamma-secretase inhibitor LY-411,575 inhibits beta-amyloid peptide production and alters lymphopoiesis and intestinal cell differentiation. J Biol Chem, 2004, 279(13): 12876-12882. |
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Cas No. | 209984-57-6 | SDF | Download SDF |
Synonyms | N/A | ||
Chemical Name | (2S)-2-[[(2S)-2-(3,5-difluorophenyl)-2-hydroxyacetyl]amino]-N-[(7S)-5-methyl-6-oxo-7H-benzo[d][1]benzazepin-7-yl]propanamide | ||
Canonical SMILES | CC(C(=O)NC1C2=CC=CC=C2C3=CC=CC=C3N(C1=O)C)NC(=O)C(C4=CC(=CC(=C4)F)F)O | ||
Formula | C26H23F2N3O4 | M.Wt | 479.48 |
Solubility | ≥23.85 mg/mL in DMSO, ≥98.4 mg/mL in EtOH with ultrasonic, <2.54 mg/ml="" in="" h2o="">2.54> | Storage | Store at -20°C |
Physical Appearance | A solid | Shipping Condition | Evaluation sample solution : ship with blue ice.All other available size:ship with RT , or blue ice upon request |
General tips | For obtaining a higher solubility , please warm the tube at 37 ℃ and shake it in the ultrasonic bath for a while.Stock solution can be stored below -20℃ for several months. |
LY-411575 is a potent inhibitor of γ-secretase with IC50 value of 0.078 nM in membrane assay.[1]γ-Secertase is one of intramenbrane-cleaving aspartyl protease which cleaves many type-I membrane proteins and many of them have important biological functions. γ-Secretase is a multi-subunit protease and it contains presenilin, nicastrin, APH-1(Anterior Pharynx- defective 1) and PEN-2. Presenilin contains Asp258 and Asp385 embedded in the sixth and seventh transmembrane domain and forms the active site. The feature of being a membrane integrated protease complex makes it difficult to be purified as well as studying its mechanism.γ-secretase is responsible for the generation Aβfrom the amyloid precursor protein.γ-Secertase has been considered as an important drug target for Alzheimer"s disease.γ-Secertase also is responsible for Notch processing which is related to cancer such as leukemia.[2]LY-411,575 significantly inhibits theγ-secretase activity in vitro. LY-411,575 inhibits the production of Aβproduction with IC50 value of 0.078 nM in membrane-assay and 0.082 nM in cell-basedγ-secretase assays, respectively. LY-411,575 also affects the Notch pathway by inhibiting Notch S3 cleavage with IC50 value of 0.39 nM.[1] LY-411,575 treatment also significantly inhibited the Notch pathway by inhibiting the γ-secretase activity luciferase activity in primary KS cells. LY-411,575 also induced the apoptosis though Notch pathway inhibition in primary and immortalized Kaposi"s sarcoma (KS) tumor cells.[2]LY-411,575 decreases the levels of brain and plasma Aβ40 and -42 at 10 mg/kg oral doses.[1] LY-411,575 also decreases cortical Aβ40 levels in transgenic CRND8 mice with ED50 ≈ 0.6 mg/kg. LY-411,575 also induced significantly intestinal goblet cell hyperplasia and thymus atrophy by inhibiting Notch signaling pathway at higher doses in vivo.[3]References: 1.Wong GT, Manfra D, Poulet FM, Zhang Q, Josien H, Bara T, Engstrom L, Pinzon-Ortiz M, Fine JS, Lee HJ et al: Chronic treatment with the gamma-secretase inhibitor LY-411,575 inhibits beta-amyloid peptide production and alters lymphopoiesis and intestinal cell differentiation. J Biol Chem 2004, 279(13):12876-12882.2.Curry CL, Reed LL, Golde TE, Miele L, Nickoloff BJ, Foreman KE: Gamma secretase inhibitor blocks Notch activation and induces apoptosis in Kaposi"s sarcoma tumor cells. Oncogene 2005, 24(42):6333-6344.3.Hyde LA, McHugh NA, Chen J, Zhang Q, Manfra D, Nomeir AA, Josien H, Bara T, Clader JW, Zhang L et al: Studies to investigate the in vivo therapeutic window of the gamma-secretase inhibitor N2-[(2S)-2-(3,5-difluorophenyl)-2-hydroxyethanoyl]-N1-[(7S)-5-methyl-6-oxo-6,7-di hydro-5H-dibenzo[b,d]azepin-7-yl]-L-alaninamide (LY411,575) in the CRND8 mouse. J Pharmacol Exp Ther 2006, 319(3):1133-1143.
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荧光定量PCR数据如下表,想通过GraphPadPrism5软件构建柱状图,但不知道是将2-△△Ct数据导入GraphPadPrism5软件,还是将Mean和SD导入GraphPadPrism5软件中。请大神赐教。另外Mean和SD是求的△△Ct的还是2-△△Ct的?
半定量反转录-聚合酶链反应(semi-quantitative reverse transcription and polymerase Chain reaction ,SqRT-PCR)是近年来常用的一种简捷、特异的定量RNA测定方法,通过mRNA反转录成cDNA,再进行PCR扩增,并测定PCR产物的数量,可以推测样品中特异mRNA的相对数量。
定量RT-PCR(quantitative reverse transcription and polymerase Chain reaction ,qRT-PCR)是在用一步法或两步法,在PCR反应体系中加入荧光基团,利用荧光信号积累实时监测整个PCR进程,最后通过标准曲线对未知模板进行定量分析的方法。
半定量RT-PCR需要跑电泳,根据条带亮度的强弱来判断模板拷贝数的高低或者是表达量的高低,而定量RT-PCR则无需电泳可以实时监测整个PCR的全程并且由给出的Ct值及Standard Curve来判断gene拷贝数的高低。
有大神做过人外周血全血miRNA的提取及后续PCR实验的吗?目前课题实验遇到瓶颈,求助大神!!!是否miRNA的PCR验证在全血做不出?必须要用血浆或者血清?目前血浆、血清也有部分样本。。。。有大神指条明路吗?
另外还有几个问题。就是跑出来的结果RFU居然有负值,这是怎么回事呢?几乎在0附近,但是在溶解曲线求导前的那个曲线。RFU又是从2400+开始下降的,这是为什么呢?
一般说来,应选用口碑较好的经过大量客户实验验证的荧光定量PCR试剂盒,因为只有选对了试剂盒才能够做成漂亮的实验结果来,特别是对于一些珍贵的样品更不能因为节约一些蝇头小利而糟蹋了样品本身。OneShineSybrGreenpreMix试剂盒在研发之初就考虑到了一般科研环境中方方面面的影响,例如我们就光照强度、光照时间对本产品的影响做了严苛的实验论证,得到1ml本产品盛装于1.5ml离心管中放置在2000Lux光照强度下可以保存12h,超过12h产品性能则会下降。
OneShineSybrGreenpreMix这款试剂盒除了价格适中外,还主要有以下优点:
1、适用于RealTimePCR反应,可以快速、准确地对目的基因进行定量检测。
2、在2×OneShineSybrGreenpreMix中,预先混有SybrGreen,PCR体系配制时只需入模板、引物、ddH2O即可,操作方便快捷。
3、含有更耐高温并持续稳定的DNAPolymerase,可以维持更多的循环数,提供了选择循环数更大的空间。更大的Ct值选择范围就意味着可以支持微量目的基因检测,检测范围fM-nM。
4、反应的Buffer更适合,使反应的基线更水平,无荧光污染信号。使线性期更陡峭,避免了Ct值的无效浮动。使平台期更平直。也确保了引物与模板能够更特异地结合。
5、SybrGreen更耐高温更足量,确保只要扩增反应在进行,目的DNA双链在增加,就有线性强度的荧光产生。
6、dNTPs更足量,确保反应有更高的平台期荧光信号。
7、Mg2+浓度更适合,确保线性期有爆发式的目的DNA片段合成,提高了线性期反应体系的扩增系数和线性期的长度。
8、更高的仪器适配性,SybrGreen染料可以被各大主流仪器的激发光所激发,产生明亮的荧光信号,有利于仪器对信号的捕捉和处理。
我们的产品与市面上目前常见的一种产品相比较有如下区别:
A公司的产品没有熔解温度,或者熔解温度太高。但是A公司的产品Ct值出现略早,这是因为A公司使用了一种小分子量的DNA聚合酶,这种酶与模板和引物的结合较快和较松散,但是扩增的效率和扩增的特异性就不能保证了,类似于温水煮青蛙不温不火的;这种酶还有一个弱点就是不耐冻融。我们使用的是Taq酶,是一种分子量较大的DNA聚合酶,这种酶与模板和引物结合比较慢,但是慢工出细活,这种酶的特异性较好,一旦激活便呈现爆发式的扩增,类似于不鸣则已一鸣惊人。我们产品熔解温度是A公司不可比拟的,线性范围和斜率也是A公司不可比拟的。详见下图:
1.模板提取(一般为RNA):Trizol、氯仿、异丙醇、无水乙醇、DEPC处理水
2.模板浓度测定:分光光度计或NanoDrop
3.逆转录:逆转录试剂盒(或者一步法试剂盒),这一步可以用普通PCR做,也可以用水域做。
4.荧光定量PCR试剂:通常有用SYBR Green Mix做的,但是这里建议你用EvaGreen做,灵敏度和平行性都要好于SYBR Green,并且如果你那是ABI或者Stratagene的PCR如果用SYBR Green还需要加一步Rox很麻烦。
5.其他:除了以上的那些还需要离心管、PCR管或板(Axygen反应比较好)、移液枪等,暂时就想到这么多。
跑到还有半小时的时候,跳出一个对话框:“analysiscannotproceed:notenoughsamplesdefined”,最后做出来的结果有扩增曲线但是没有熔解曲线,请各位大神帮忙解释一下。
如果要测量细胞表面2个受体的比率,最好用流式细胞仪来测量
在另外的一组管子中,加入已知拷贝数的DNA同时扩增,每个管子中加入的数量是不同的,但是从最小数目到最大数目的这个范围涵盖了样本中DNA拷贝数。
PCR反应完成后,把已知拷贝数DNA的量和PCR荧光Ct值制成标准曲线,再把待测样本的CT值和该标准曲线比对,就可以得到样本中的起始拷贝数了