Cell Signaling Technology

Product Pathways - Apoptosis

PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit #11874

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No. Size Price
11874S 1 Kit ( 96 assays ) ¥6,345.00 现货查询 购买询价
Product Includes Volume Solution Color
Bcl-2 Mouse Detection Ab 11 ml Green
ELISA Sample Diluent 25 ml Blue
STOP Solution #7002 11 ml Colorless
TMB Substrate #7004 11 ml Colorless
Anti-mouse IgG, HRP-linked Antibody 11 ml Red
ELISA Wash Buffer (20X) 25 ml Colorless
Cell Lysis Buffer (10X) #9803 15 ml Yellowish
Phospho-Bcl-2(Ser70) Rabbit Ab Coated Microwells 96 tests
Sealing Tape 2 sheets

Specificity / Sensitivity

The PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit recognizes endogenous levels of Bcl-2 protein when phosphorylated at Ser70 as shown in Figure 1. Kit sensitivity is shown in Figure 2. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

The PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit能够检测内源的磷酸化Bcl-2(Ser70)蛋白,如图1所示。试剂盒灵敏度如图2所示。经内部测试验证,本试剂盒能够检测指定物种的蛋白,同时也可以检测其他物种的同源蛋白。

Description

The PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Bcl-2 when phosphorylated at Ser70. A Phospho-Bcl-2 (Ser70) Rabbit mAb has been coated onto the microwells. After incubation with cell lysates, phosphorylated Bcl-2 protein is captured by the coated antibody. Following extensive washing, a Bcl-2 Mouse Detection mAb is added to detect the captured phospho-Bcl-2 (Ser70) protein. Anti-mouse IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of absorbance for the developed color is proportional to the quantity of Bcl-2 phosphorylated at Ser70. 
 Antibodies in kit are custom formulations specific to kit.

PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit采用了固相夹心酶联免疫吸附方法(ELISA)检测内源的Ser70磷酸化的Bcl-2蛋白。微孔中包被了Phospho-Bcl-2 (Ser70) Rabbit mAb 兔单抗。细胞裂解液孵育后,磷酸化的Bcl-2蛋白被包被的抗体捕获。充分洗涤后,添加Bcl-2 Mouse Detection mAb检测捕获的Ser70磷酸化的Bcl-2蛋白。再用Anti-mouse IgG和HRP-linked Antibody识别结合的抗体。加上HRP底物,TMB显色。测得的吸光值与Ser70磷酸化Bcl-2蛋白的量成正比。试剂盒中的抗体为该试剂盒所特制。

ELISA - Western correlation

ELISA - Western correlation

Figure 1. Treatment of Jurkat cells with Paclitaxel stimulates phosphorylation of Bcl-2 at Ser70, as detected by the PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit, but does not affect the levels of total Bcl-2 detected by PathScan® Total Bcl-2 Sandwich ELISA Kit #12030. Jurkat cells were untreated or treated with λ phosphatase or Paclitaxel #9807 (1 mM, 20 hr, 37ºC). The absorbance readings at 450 nm are shown in the top figure, while the corresponding western blots using Phospho-Bcl-2 (Ser70) (5H2) Rabbit mAb #2827 (left panel) or Bcl-2 (D55G8) Rabbit mAb (Human Specific) #4223 (right panel) are shown in the bottom figure.

图1. PathScan® Phospho-Bcl-2 (Ser70) Sandwich ELISA Kit检测经处理的Jurkat细胞,Paclitaxel刺激Bcl-2蛋白的Ser70磷酸化,但是不影响PathScan® Total Bcl-2 Sandwich ELISA Kit #12030检测Bcl-2总蛋白。Jurkat细胞经未处理或λ磷酸酶或Paclitaxel #9807 (1 mM, 20 hr, 37ºC)处理。450 nm吸光值见上图,相应的western blots 分析见下图,所用抗体为Phospho-Bcl-2 (Ser70) (5H2) Rabbit mAb 兔单抗#2827 (左)和 Bcl-2 (D55G8) Rabbit mAb (Human Specific) 兔单抗#4223(右)。

ELISA

ELISA

Figure 2. The relationship between protein concentration of lysate from Jurkat cells, untreated or treated with λ phosphatase or Paclitaxel #9807 (1 mM, 20 hr, 37ºC), and the absorbance at 450 nm is shown.

图2. 如图所示为Jurkat细胞裂解液的蛋白浓度之间的关系,未经处理的或用λ磷酸酶或Paclitaxel #9807 (1 mM, 20 hr, 37ºC)处理,显示450 nm处的吸光度。

Background

Bcl-2 exerts a survival function in response to a wide range of apoptotic stimuli through inhibition of mitochondrial cytochrome c release (1). It has been implicated in modulating mitochondrial calcium homeostasis and proton flux (2). Several phosphorylation sites have been identified within Bcl-2 including Thr56, Ser70, Thr74, and Ser87 (3). It has been suggested that these phosphorylation sites may be targets of the ASK1/MKK7/JNK1 pathway and that phosphorylation of Bcl-2 may be a marker for mitotic events (4,5). Mutation of Bcl-2 at Thr56 or Ser87 inhibits its anti-apoptotic activity during glucocorticoid-induced apoptosis of T lymphocytes (6). Interleukin-3 and JNK-induced Bcl-2 phosphorylation at Ser70 may be required for its enhanced anti-apoptotic functions (7).

Bcl-2 通过抑制线粒体细胞色素C的释放而对一系列的凋亡因素做出响应,从而发挥其促进细胞存活的功能(1)。这提示其在调节线粒体钙离子的动态平衡和质子流方面发挥着作用(2)。在Bcl-2中已经发现了几个磷酸化位点,包括Thr56, Ser70, Thr74 和 Ser87 (3)。这些磷酸化位点可能是ASK1/MKK7/JNK1通路的靶点,并且Bcl-2的磷酸化可能是有丝分裂活动的一个标志(4,5)。在T淋巴细胞糖(肾上腺)皮质激素诱导的凋亡过程中,Bcl-2在Thr56或者Ser87位点的突变能够抑制其抗凋亡的活性(6)。白细胞介素3 和JNK诱导的Bcl-2在Ser70 位点的磷酸化可能是其增强抗凋亡作用所必需的(7)。

  1. Murphy, K.M. et al. (2000) Cell Death Differ 7, 102-11.
  2. Zhu, L. et al. (1999) J Biol Chem 274, 33267-73.
  3. Maundrell, K. et al. (1997) J Biol Chem 272, 25238-42.
  4. Yamamoto, K. et al. (1999) Mol Cell Biol 19, 8469-78.
  5. Ling, Y.H. et al. (1998) J Biol Chem 273, 18984-91.
  6. Huang, S.T. and Cidlowski, J.A. (2002) FASEB J 16, 825-32.
  7. Deng, X. et al. (2001) J Biol Chem 276, 23681-8.

Application References

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Protocols


For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

PathScan is a trademark of Cell Signaling Technology, Inc.

Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.

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