Quantitative detection of red blood cell antibody-mediated complement activation
10.13303/j.cjbt.issn.1004-549x.2022.09.025
- VernacularTitle:红细胞抗体介导的补体活化的定量检测
- Author:
Zhongying WANG
1
;
Jian LI
2
;
Fengyong ZHAO
1
;
Chenrui QIAN
1
;
Wei SHEN
1
;
Liangfeng FAN
1
;
Sha JIN
1
;
Jiewei ZHENG
1
;
Yuyu ZHANG
1
;
Dong XIANG
1
Author Information
1. Shanghai Blood Center, Shanghai 200051, China
2. Shanghai United Family Hospital Co., Ltd
- Publication Type:Journal Article
- Keywords:
complement activation;
red blood cell antibody;
flow cytometry;
spectrophotometry
- From:
Chinese Journal of Blood Transfusion
2022;35(9):982-985
- CountryChina
- Language:Chinese
-
Abstract:
【Objective】 To construct an in-vitro model of erythrocyte antibody-mediated complement activation, and establish quantitative detection methods based on flow cytometry and spectrophotometry, so as to explore the correlation of anti-body titers and complement activation speed, and provide a methodological basis for studying the adverse transfusion reactions of anti-body mediated complement hemolysis. 【Methods】 Mouse monoclonal antibody that recognized human C3b and fluorescent secondary antibody were used to label C3b fragments on erythrocytes, and the deposition of C3b fragments after complement activation was detected by flow cytometry. The absorbance at 540 nm of the supernatant in the complement activation reaction system was measured by spectrophotometry as the amount of hemoglobin released was related to the absorbance. 【Results】 The complement activation system was constructed according to the ratio of 3% red blood cell suspension (mixed for 6 people) 1∶anti-Tja 1∶complement 2. The repeatability was good (P value>0.05) as different red blood cell mixtures had been used to repeat the detection reaction system. When using 32×, 64× and 128× dilutions of anti-Tja mediated complement activation, the deposition of C3b fragments has been detected by flow cytometry at 30 s, 1 min and 2 min, respectively, and MFI peaked at 5 min, 10 min and 30 min, respectively. No obvious hemolysis has been observed within 1.5 h. 【Conclusion】 In vitro model of anti-Tja-mediated complement activation demonstrates the speed of complement activation is related to the concentration of antibody. At a certain antibody concentration, the speed of complement activation has been slowed down, and no obvious hemolysis observed.
