论文部分内容阅读
本文介绍检测免疫球蛋白阳性细胞(IpCS)的一种敏感ELISA技术。作者用5×10~3/mlTNP—SRBC 0.5ml免疫6—8周龄CBA系雌性小鼠,7天后取其脾(Sp)和肠集合淋巴结(Pp)分离淋巴细胞。用组织培养微量滴定板进行培养,分别加入LPS、LPS+ConA、SRBS进行刺激,培养7天后,做细胞ELISA检查IPC,并检查培养上清液Ig总量。细胞ELI-SA方法如下: 1、细胞培养7天后,取其上清液,保存于-20℃,每孔中加含0.05%吐温20的磷酸盐缓冲液(PBS/T)200±μl。在微量滴定板振荡器上振动30秒。2、然后用平板离心器800rpm离心10分钟,弃上清液后,37℃干燥30分钟。3、每孔加甲醇50μl,置室温固定5分钟,除去多余的甲醇,空气干燥微量板。
This article describes a sensitive ELISA technique for detecting immunoglobulin-positive cells (IpCS). The mice were immunized with 0.5ml 5 × 10-3 / ml TNF-SRBC for 6-8 weeks old CBA female mice. After 7 days, the spleen (Sp) and intestinal collecting lymph nodes (Pp) were used to isolate lymphocytes. Tissue culture microtiter plates were used for the culture. LPS, LPS + ConA and SRBS were added to the culture for 7 days. Cell culture was used to test the IPC and the amount of Ig in culture supernatants was examined. The cell ELI-SA method is as follows: 1. After the cells were cultured for 7 days, the supernatant was taken and stored at -20 ° C. 200 ± μl of phosphate buffered saline (PBS / T) containing 0.05% Tween 20 was added to each well. Vibration for 30 seconds on a microtiter plate shaker. 2. Centrifuge at 800rpm for 10 minutes using a plate centrifuge, discard the supernatant, and then dry at 37 ° C for 30 minutes. 3, add 50μl of methanol to each well, set at room temperature for 5 minutes, remove excess methanol and air dry the microplate.