P2z/P2x7受体检测的研究

Detection of P2z/P2x7 Receptors

目的 检测白血病细胞的 P2 z/ P2 x7受体。方法 将 T淋巴细胞白血病细胞株 (CEM)、急性单核细胞白血病细胞株 (THP- 1)、M2 型白血病细胞株 (HL - 6 0 )、M3型白血病细胞株 (NB4 )、巨核细胞病细胞株 (Dam i/Dam)和伯基特淋巴瘤细胞株 (Raji)共六株白血病细胞株制成 2× 10 7/ m l的细胞悬液 ,分别加入 P2 z/ P2 x7受体的激动剂 ATP和 /或特异性抑制剂 KN- 6 2 ,测定加入前后细胞内荧光强度变化 ,同时以 P2 z/ P2 x7受体多克隆抗体免疫组化 ABC法鉴定 P2 z/ P2 x7受体。结果 10 .4 mm ol/ L 的 ATP对于 THP- 1、NB4和 HL- 6 0细胞株钙离子通透性有较强的激活作用 ,其激活率分别为 5 4 .91%、39.4 5 %和 30 .4 6 % ;ATP对 CEM、Dam和 Raji细胞株的激活作用很弱 ,激活率分别为 10 .4 35 %、13.2 15 %和 2 .6 7% ;KN- 6 2对 THP- 1有很强的抑制作用 (抑制率达 88.74 % ) ,而对 NB4和 HL- 6 0的抑制作用很弱 (抑制率仅为 15 .81%和 12 .5 0 % ) ,对 Raji细胞株没有抑制作用 ;2免疫组化ABC法显示各对照细胞株均成阴性 ,而白血病细胞株均成阳性。结论 THP- 1细胞含 P2 z/ P2 x7受体 ,NB4和 HL-6 0含 P2受体其它亚型的细胞株 ,CEM、Raji和 Dam细胞株不含 P2 z/ P2 x7受体 ;免疫组化 ABC法使用的多克隆抗?

Objective To detect the P2z/P2x7 receptors of human leukocytes. Methods Re suspending different kinds of leukemic cells, including CEM, THP 1, HL 60, NB4, Dami/Damin and Raji cells in buffer; adjusting their concentration to 2×10 7/ml, and then detecting the P2z/P2x7 receptors respectively. Results ① It was found that 0.4 mmol/L ATP enhanced obviously the fluorenscence strength of THP 1, NB4 and HL 60 cells, the activity rates were 54.91%, 39.45% and 30.46%, respectively. But for CEM, Dam and Raji cells, the activity rates were as low as 10.43%, 13.21% and 2.67%, respectively. KN 62 inhibited obviously the fluorescence strength of THP 1 cells, the inhibition rate reached 88.74%, but for NB4 and HL 60 cells the inhibition rates were only 15.81% and 12.50% respectively, and Raji cells were not affected by KN 62. ② All the leukemic cells detected with polyclonal antibody were found with positive result. Conclusion THP 1 cells have P2z/P2x7 receptors.The receptors in NB4 and HL 60 belong to P2 receptor family, but they are not P2z/P2x7 receptors. The polyclonal antibody used in immunohistochemical ABC method is not specific to P2z/P2x7 receptors.