一种低成本、低本底的免疫细胞化学反应方法

AN IMMUNOCYTOCHEMICAL METHOD WITH LOW BACKGROUND STAINING AND LOW EXPERIMENTAL COST

为了探讨一种适合培养细胞的低本底、低成本免疫细胞化学方法 ,本研究将低浓度的 Triton X-10 0加入抗体稀释液和洗液中 ,观察了其对 ABC反应时的抗体和 ABC的使用浓度和反应效果的影响。结果证明 ,在抗体稀释液和洗液中加入 0 .1%Triton X-10 0可以增强细胞对抗体的通透性 ,在不减弱阳性信号的状况下使 -抗的使用浓度明显降低 ,使生物素化二抗和 ABC的使用浓度达到 1/ 10 0 0～ 1/ 40 0 0 ,并大大减弱染色本底。本研究结果提示 ,在这一稀释度明显低于目前试剂盒建议的使用浓度。在充分显示阳性信号的前提下 ,使抗体的使用浓度和量大幅度减少 。

To explore an immunocytochemical method with low background staining and low experimental cost that is suitable for cultured cells, the effects of application of low concentration of Triton X-100 to antibody diluting solution and washing solution on the concentrations of antibody and ABC and the staining result in immunocytochemical ABC mehtod were investigated in the present study. The results showed that addition of 0.1% Triton X-100 to antibody diluting solution and washing solution significantly increased the permeability of cell membrane to antibody and decreased the dilutions of biotinylated second antibody and ABC to 1/1000～1/4000 without compromising positive signal, and simultaneously reduced the background staining. The results indicate that under the circumstances of fully displaying positive signal, the usage of antibody can be greatly decreased by application of low concentration of Triton X-100 to antibody diluting solution and washing solution, thereby realizing the purpose of reducing staining background and experimental cost. (Figures A～H on plate 35)