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Cignal Lenti Reporter Assays

用于灵敏检测各种哺乳动物细胞的信号活性

  • 可转导各种哺乳动物细胞
  • 转导未分化细胞、干细胞和分化细胞
  • 用于瞬转实验或研发稳定细胞株的报告因子
  • 可即用于转导的慢病毒,无需额外制备过程

Cignal Lenti Reporter Assays为可即用于转导的慢病毒颗粒,可检测各种哺乳动物细胞的信号活性。

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41
产品 Product no. 货号 目录价:
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-001L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-002G varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-002L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-003L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-007L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-008L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-009L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-010G varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-010L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-011G varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-011L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-012L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-013G varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-013L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-014G varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-014L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-015L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-016L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-017L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-018G varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-018L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-1036L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-2020L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-2033L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-3027L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-3030L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-4024L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-4037L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-4040L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-5021L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-5034L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-6028L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-6031L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-6044L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-7025L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-7041L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-8019L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-8022L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-9029L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
show details
336851 CLS-9032L varies
250 µl ready-to-transduce lentivirus reporter at an approximate titer of 1 x 107 transduction units (TU) per ml and is sufficient to transduce ~2.5 x 106 cells at a multiplicity of infection (MOI) of 1 TU/cell
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336851 CLS-9045L varies
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Cignal Lenti Reporter Assays适用于分子生物学应用。该产品不适用于疾病的诊断、预防或治疗。


0
确定人类原代细胞的通路活性。
Cignal Lenti NFAT Reporter Assay用于确定PKA/Ca2+信号通路的活性。Cignal Lenti NFAT Reporter Assay (luc) (4 x 105 TU)和Cignal Lenti Renilla Control (luc) (1 x 105 TU)共转导大约10,000个正常人类肺动脉平滑肌细胞(PASMC)。转导48小时后,将培养基换成分析培养基。转导54小时后,细胞用10 ng/ml PMA和0.5 µM钙离子载体处理18小时。随后进行双荧光素酶分析,启动子的活性用Renilla作为内参标准化后的任意值表示。每个实验均重复三次,标准差如图所示。
1
大鼠脑胶质瘤细胞的Notch信号传导活性。
Cignal Lenti RBP-Jk Reporter Assay用于检测C6(大鼠脑胶质瘤)细胞中Notch信号传导的活性。Cignal Lenti RBP-Jk Reporter Assay (luc) (2 x 105 TU)和Cignal Lenti Renilla Control (luc; 2 x 104 TU)共转导大约20,000个C6细胞。转导24小时后,将培养基换成完全培养基。转导48小时后,用100 MOI表达激活状态Notch1 (Ad-NICD)的重组腺病毒或100 MOI表达GFP (Ad-GFP)的重组腺病毒感染细胞。感染18小时后,进行双荧光素酶分析,启动子的活性用Renilla作为内参标准化后的任意值表示。每个实验均重复三次,标准差如图所示。
2
测量NFκB信号通路活性。
Cignal Lenti NFκB Reporter Assay用于检测胸腺细胞(D1;小鼠T细胞白血病细胞)中NFκB信号通路的活性。Cignal Lenti NFκB Reporter Assay (luc) (2.5 x 105 TU)转导大约10,000个D1细胞。转导48小时后,细胞用20 ng/ml重组鼠TNFα (hTNFα)蛋白处理18小时。随后进行荧光素酶分析,启动子的活性用任意值表示。每个实验均重复三次,标准差如图所示。
3
采用双荧光素酶系统的优势。
Cignal Lenti HIF-1 Reporter (luc) (1.5 x 105 TU)转导大约15,000个HepG2细胞(转导之前24小时将细胞种到96孔板中,7,500个细胞/孔),使用Cignal Lenti Renilla对照或不使用均可。转导42小时后,将培养基换成分析培养基(Opti-MEM + 0.5% FBS + 0.1mM NEAA + 1mM丙酮酸钠+ 100 U/ml青霉素+ 100 µg/ml链霉素)。转导48小时后,细胞用300 µM的CoCl2处理18小时。随后进行荧光素酶或双荧光素酶分析,启动子的活性用Renilla作为内参标准化后的任意值表示。每个实验均重复三次,标准差如图所示。利用双荧光素酶体系(共转导Cignal Lenti Renilla Control和Cignal Lenti Reporter Assay (luc))提高了缺氧信号活性双重诱导的变化,同时萤火虫荧光素酶的活性用海肾荧光素酶标准化后,修正了由于加药处理后细胞死亡的影响。
4
建立稳定的细胞株。
Cignal Lenti NFkB Reporter Assay (luc)用于建立一个NFkB信号通路敏感的细胞株,用来研究NFkB信号转导通路。NFkB敏感细胞株的建立:HEK-293细胞转导Cignal Lenti NFkB Reporter Assay (luc),筛选染色体中稳定整合病毒并且对NFkB信号通路的激活反应强烈的克隆群。在该细胞株的第1、5、10和15个传代后,使用10 ng/ml的重组hTNFα来确定稳转HEK-293 NFkB敏感细胞株是否建立成功。即使在培养细胞2个月之后,使用hTNFα激活NFkB通路,报告基因的表达仍有100倍的提高。
5
Cignal Lenti Reporter Assay procedure.
6
Cignal Lenti Reporter Assay.
7
Cignal Lenti Reporter Assays.
Performance
Cignal Lenti Reporter Assays非常适合研究原代细胞、干细胞和难转染细胞株的细胞信号(参见"Determination of pathway activity in human primary cells"、"Notch signaling activity in rat glioma cells"、"Measurement of NFkB pathway activity"和"Advantage of using dual luciferase system")。还可用于制备稳定细胞株(参见"Stable cell line generation")和siRNAs的高通量筛选。
Principle

Cignal Lenti Reporter Assays独特的整合了转录因子报告基因技术和慢病毒技术。Cignal Lenti Reporter Assays含有多拷贝的特定转录因子结合部位和基本启动子元件,可表达报告基因(萤火虫荧光素酶或GFP)。

慢病毒是将含报告基因的载体转入大部分哺乳动物细胞(包括未分化细胞)的最有效工具。转导的慢病毒报告基因载体与细胞基因组DNA整合,可稳定、长期表达报告基因。

这些报告基因是功能基因组学和药物研发的有力工具,可检测通路活性。当候选药物、基因敲除(使用siRNA)、过表达(使用表达载体)或多肽激活或抑制通路时,荧光素酶或GFP报告基因活性可被调控,并能被快速、定量检测。

安全性

Cignal Lenti Reporter Assays是转导即用型、不能复制的、基于HIV的VSV-G假慢病毒颗粒(参见"Cignal Lenti Reporter Assay")。Cignal慢病毒颗粒使用安全。推荐按照Risk Group Level 2 (RGL-2)生物进行处理。按照所有发行的RGL-2准则进行操作和废物净化。建造BSL-2工作环境的详细要求可参见U.S. Department of Health and Human Services出版的Biosafety in Microbiological and Biomedical Laboratories,第4版。

这些载体经设计改造后具有以下生物安全特征:

3' LTR的U3部分的启动子/增强子区域被去除,保证慢病毒载体转导并与目标细胞的基因组DNA整合后自我失活
Cignal Lenti Reporter Assay和质粒表达的包装蛋白无明显的同源区,减少重组的可能性
没有任何HIV-1基因(gag、pol和rev)可在转导细胞中表达,因为它们是由不含包装信号的包装质粒表达的。因此,慢病毒颗粒无法复制
Cignal Lenti Reporter Assay不含病毒性基因(vpr、vif、vpu和nef) 
Cignal Lenti Reporter Assays的特征
特征 功能
RSV-5' LTR:融合肉瘤病毒(RSV)增强子/启动子-U5长末端重复 包装病毒和反转录病毒mRNA
Psi:包装信号 允许病毒包装
RRE:病毒响应元素 参与病毒转录物包装
Cppt:中心聚嘌呤区 参与核转移和整合转导的病毒基
报告基因(萤火虫荧光素酶或GFP) 能够定量转录
hPGK:人磷酸甘油酸激酶真核细胞启动子 可高水平表达哺乳动物筛选标记(嘌呤霉素)
PuroR:嘌呤霉素抗性基因 用于哺乳动物筛选
SIN/3' LTR:3'自动失活长末端重复 经修饰的3' LTR可包装病毒,但自动失活5' LTR,确保生物安全;该元素也包含一个多聚腺苷酸化信号,用于有效终止转录
f1 ori:f1复制起点 在真核细胞中游离复制质粒
AmpR:氨苄青霉素抗性基因 E. coli中筛选质粒
TRE:转录响应元素 通过特有的转录因子调控报告基因的表达
TATA阅读框 较小的启动子
Procedure

Cignal Lenti Reporter Assays可直接转导,无需构建或扩增慢病毒。这些载体非常适合对难转染细胞瞬时转导或制备通路传感细胞株。

 难转染细胞株的瞬时通路调控研究

使用Cignal Lenti Reporter Assay转导目标细胞。细胞通常培养24–48小时,以确保慢病毒整合。再用相应的生物试剂处理(siRNA、shRNA、化合物、病毒表达载体、蛋白或多肽)。根据处理条件,处理后18–36小时进行报告基因检测(萤火虫荧光素酶或GFP)(参见"Cignal Lenti Reporter Assay procedure")。

制备稳定的通路传感细胞株

使用Cignal Lenti Reporter Assay转导目标细胞。转导后,细胞用嘌呤霉素筛选,形成转导细胞的纯细胞群落。如果必要,可进行单细胞克隆,进而分离通路传感细胞株。这些通路传感细胞株可作为有用的细胞分析平台,用于后续筛选和机理研究。

Applications

Cignal Lenti Reporter Assays适用于功能基因组学和药物研发,可检测各种哺乳动物细胞的信号活性。非常适合检测siRNAs、蛋白和小分子化合物的生物效应。

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产品介绍与指南 (1)
For measurement of signaling pathways in any mammalian cell
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试剂盒操作手册 (1)
For lentiviral-based cell signaling activity assays
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