In vitro method establishment for detecting monocyte phagocytosis of sensitized RBCs by flow cytometry
10.13303/j.cjbt.issn.1004-549x.2022.08.008
- VernacularTitle:流式细胞术检测单核细胞体外吞噬致敏红细胞实验方法的建立
- Author:
Siying ZHU
1
;
Yanli JI
1
;
Zhen WANG
1
;
Dawei CHEN
1
;
Zhijian LIAO
1
;
Chunyan MO
1
;
Jizhi WEN
1
;
Guangping LUO
1
;
Ling WEI
1
Author Information
1. Institute of Clinical Blood Transfusion, Guangzhou Blood Center, Guangzhou 510095, China
- Publication Type:Journal Article
- Keywords:
monocytes;
phagocytosis;
flow cytometry;
monocyte monolayer assay (MMA)
- From:
Chinese Journal of Blood Transfusion
2022;35(8):814-816
- CountryChina
- Language:Chinese
-
Abstract:
【Objective】 To establish an experimental method for detecting phagocytosis of sensitized red blood cells in vitro by flow cytometry. 【Methods】 Mononuclear cells were isolated from the peripheral blood of blood donors and cultured in a cell incubator for 1 hour, and then adherent monocytes were isolated and obtained. Dib-positive red blood cells (RBCs) were labeled with PKH26 and then sensitized with IgG anti-Dib. The sensitized RBCs were added to monocytes for in vitro phagocytosis assay. Monocytes were labeled with FITC anti-human CD14, then phagocytosis was measured by flow cytometry, and the phagocytic efficiency was calculated. The method was used to detect the phagocytic efficiency of monocytes on human IgG anti-D sensitized RBCs with different titers. 【Results】 The phagocytic efficiency of monocytes was averaged at 5% (1.2%~7.6%, SD 3.30) versus 81% (71.4%~92.7%, SD 8.65) in the negative versus positive control group, respectively. Phagocytic activity of monocytes mediated by anti-D was correlated with the antibody titer. The phagocytosis efficiency was within 10% when the antibody titer was lower than 32 and increased sharply when the titer was between 32 to 128, it entered a plateau and stabilized at 80% at the titer above 256. 【Conclusion】 A detection platform for detecting phagocytosis-sensitized RBCs in vitro by flow cytometry has been successfully established. It can be used to assess the clinical significance of red blood cell allotype or autologous IgG antibodies.
