| 货号 | 7938C |
| 描述 | The PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of eIF4E when phosphorylated at Ser209. An eIF4E mouse antibody has been coated onto the microwells. After incubation with cell lysates, eIF4E (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, an eIF4E rabbit detection antibody is added to the captured phospho and nonphospho eIF4E protein. Anti-rabbit IgG, HRP-linked Antibody #7074 is then used to recognize the bound detection antibody. HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of phospho-eIF4E (Ser209). Antibodies in kit are custom formulations specific to kit.该PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit是一个固相夹心酶联免疫吸附法（ELISA），其能检测在Ser209位点磷酸化的内源性eIF4E蛋白。一个eIF4E鼠源已经被包被在微孔板上。在与细胞裂解液一起孵育后，eIF4E (磷酸和非磷酸化的)蛋白被包被的抗体吸附。在充分的清洗后，加入一个eIF4E兔源检测抗体以检测已被吸附的磷酸化和未磷酸化的eIF4E蛋白。然后，Anti-rabbit IgG, HRP-linked Antibody #7074被用于识别已结合的检测抗体。加入HRP底物TMB后显色。该颜色的吸光度值与磷酸化eIF4E (Ser209)蛋白数量成正比。试剂盒里的抗体是针对该试剂盒专门定制的。 |
| 反应种属 | Human |
| 应用 | ELISA |
| 目标/特异性 | CSTs PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit #7938 detects endogenous levels of eIF4E protein when phosphorylated at Ser209. As shown in Figure 1, a significant induction of eIF4E phosphorylation at Ser209 can be detected in HeLa cells following treatment with 20% FBS using the Phospho-EIF4E (Ser209) Sandwich ELISA Kit. The levels of total eIF4E remain unchanged as shown by western analysis and by PathScan® Total eIF4E Sandwich ELISA Kit #7940 (Figure 1). This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species. |
| 供应商 | CST |
| 背景 | Eukaryotic initiation factor 4E (eIF4E) binds to the mRNA cap structure to mediate the initiation of translation (1,2). eIF4E interacts with eIF4G, a scaffold protein that promotes assembly of eIF4E and eIF4A into the eIF4F complex (2). eIF4B is thought to assist the eIF4F complex in translation initiation. Upon activation by mitogenic and/or stress stimuli mediated by Erk and p38 MAPK, Mnk1 phosphorylates eIF4E at Ser209 in vivo (3,4). Two Erk and p38 MAPK phosphorylation sites in mouse Mnk1 (Thr197 and Thr202) are essential for Mnk1 kinase activity (3). The carboxy-terminal region of eIF4G also contains serum-stimulated phosphorylation sites, including Ser1108, Ser1148, and Ser1192 (5). Phosphorylation at these sites is blocked by the PI3 kinase inhibitor LY294002 and by the FRAP/mTOR inhibitor rapamycin. |
| 存放说明 | 4C |
| 参考文献 | Sonenberg, N. et al. (1978) Proc. Natl. Acad. Sci. USA 75, 4843-4847. Gingras, A.C. et al. (1999) Annu. Rev. Biochem. 68, 913-963. Waskiewicz, A. et al. (1999) Mol. Cell. Biol. 19, 1871-1880. Pyronnet, S. et al. (1999) EMBO J. 18, 270-279. Raught, B. et al. (2000) EMBO J. 19, 434-444. |
Figure 1. Treatment of serum-starved HeLa cells with 20% FBS stimulates phosphorylation of eIF4E at Ser209, detected by the PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit #7938, but does not affect the levels of total eIF4E detected by PathScan® Total eIF4E Sandwich ELISA Kit #7940. HeLa cells (80-90% confluent) were serum-starved over night and then treated with 20% FBS at 37ºC. The absorbance readings at 450 nm are shown in the top figure, while the corresponding western blots using eIF4E Antibody #9742 (left panel) or Phospho-eIF4E Antibody (Ser209) #9741 (right panel) are shown in the bottom figure. 图1：使用20% FBS处理的血清饥饿培养的HeLa细胞刺激eIF4E蛋白在Ser209位点磷酸化，随后使用PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit #7938检测。但是这不影响通过PathScan® Total eIF4E Sandwich ELISA Kit #7940所检测的eIF4E总蛋白水平。在饥饿培养过夜后，HeLa细胞在37ºC(80-90% confluence)给予20% FBS在处理。450 nm的吸光度值显示在最上图中，使用使用eIF4E Antibody #9742 (左图)和Phospho-eIF4E Antibody (Ser209) #9741 (右图)进行的western blot分析显示在下图中。 | |
Figure 2. The relationship between the protein concentration of lysates from untreated and 20% FBS-treated HeLa cells and the absorbance at 450 nm using the PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit is shown.Figure 2. The relationship between the protein concentration of lysates from untreated and 20% FBS-treated HeLa cells and the absorbance at 450 nm using the PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit is shown. 图2：使用PathScan® Phospho-eIF4E (Ser209) Sandwich ELISA Kit，对未处理组和20% FBS处理组的HeLa细胞裂解物进行分析，相应的蛋白浓度与在450 nm吸光度值之间的关系如图所示。 |
