使用生物素-抗生物素蛋白红细胞花环法分离淋巴细胞亚群。

Separation of lymphocyte subpopulations using biotin-avidin erythrocyte rosettes.

作者信息

Levitt D, Danen R

出版信息

J Immunol Methods. 1986 May 22;89(2):207-11. doi: 10.1016/0022-1759(86)90359-5.

Abstract

The major method for isolating murine lymphocyte subpopulations involves negative selection using antibody plus complement-mediated cytolysis. We have developed an efficient rosette method for enriching murine B and T cells using biotin-conjugated antibodies and avidin-coated sheep erythrocytes. Rosetted and non-rosetted subpopulations are separated rapidly on Percoll cushions. In systems employing rabbit anti-mouse immunoglobulin or monoclonal rat anti-mouse Thy-1.2 conjugated to biotin, positively-selected cells are greater than 90% pure while negatively-depleted populations possess less than 2% contamination with unwanted cells. Recoveries from starting spleen cell populations range between 50 and 75%. This method provides an easily performed alternative for obtaining positively and negatively selected cell populations and can be used with any biotin-conjugated antibody protein.

摘要

分离小鼠淋巴细胞亚群的主要方法是使用抗体加补体介导的细胞溶解进行阴性选择。我们开发了一种高效的玫瑰花结方法，利用生物素偶联抗体和抗生物素蛋白包被的绵羊红细胞富集小鼠B细胞和T细胞。在Percoll垫层上可快速分离形成玫瑰花结和未形成玫瑰花结的亚群。在使用与生物素偶联的兔抗小鼠免疫球蛋白或单克隆大鼠抗小鼠Thy-1.2的系统中，阳性选择的细胞纯度大于90%，而阴性去除的群体中不需要的细胞污染率小于2%。起始脾细胞群体的回收率在50%至75%之间。该方法为获得阳性和阴性选择的细胞群体提供了一种易于操作的替代方法，并且可与任何生物素偶联抗体蛋白一起使用。