Efficient purification and label of anti-human beta 2-microglobulin light chain monoclonal antibody
10.3969/j.issn.1673-8225.2011.24.046
- VernacularTitle:纯化和标记抗人轻链β2m单克隆抗体的方法
- Author:
Guangping RUAN
;
Xiang YAO
;
Rongqing PANG
;
Xingming WANG
;
Ying DAI
;
Xinghua PAN
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2011;15(24):4557-4560
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND: The inoculation of hybridoma cell strain onto mouse abdominal cavity may obtain ascites containing mass antibody. Previous method to purify monoclonal antibody in ascites is complex and difficult to operate.OBJECTIVE: To prepare, purify and label anti-human leukocyte antigen (HLA)-I class molecule light chain monoclonal antibody, and to detect the expression of tumor cell surface HLA-I class molecules. METHODS: Hybridoma cells were inoculated onto the mouse abdominal cavity. Ascites containing anti-human light chain beta2-microglobulin antibody were obtained and purified with the modified caprylic acid-ammonium sulfate method. The purified monoclonal antibody was labeled with fluorescein isothiocyanate to detect peripheral blood mononuclear cells, T2 cells expressing blank HLA-A2 molecule and K562 cells surface HLA-I class molecules. The expression of HLA-I class molecules was determined by using flow cytometry and fluorescent microscopy. RESULTS AND CONCLUSION: The purified anti-human light chain beta2-microglobulin-fluorescein isothiocyanate monoclonal antibodies accounted for 96% purity. Flow cytometry results showed that, the HLA-I class molecules were highly expressed in peripheral blood mononuclear cells surface, lowly expressed in T2 cells, and not expressed in K562 cells surface. It is a simple and convenient method to purify ascites with the modified caprylic acid-ammonium sulfate method, and according prepare anti-human light chain beta2-microglobulin-fluorescein isothiocyanate. This method is effective to distinguish the levels of HLA-I class molecules expressed in various cells.
