Measurement of binding affinity between peptides and HLA-B*1301 by HLA molecular reconstitution
10.3969/j.issn.1000-484X.2024.10.024
- VernacularTitle:HLA分子重构法检测多肽与HLA-B*1301的结合力
- Author:
Shuai LIU
1
;
Mengnan YI
;
Bo JIAO
;
Yican WANG
;
Yuanyuan CHEN
;
Yufei DAI
Author Information
1. 中国疾病预防控制中心职业卫生与中毒控制所化学污染物与健康重点实验室,北京 100050
- Keywords:
HLA;
Peptides;
Molecular reconstruction;
Binding affinity
- From:
Chinese Journal of Immunology
2024;40(10):2163-2167
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish a method for detecting the binding affinity of peptides to HLA-B*1301 by HLA molecular reconstitution after weak acid treatment,and its practicality was evaluated.Methods:C1R cells overexpressing HLA-B*1301 were treated with citrate buffer of different pH for different time.After neutralization of pH,cells were resuspended in culture medium con-taining β2m and brefeldin A.Cells were incubated at 37℃with peptide(peptide group)or without peptide(blank control),after addition of anti-HLA antibody,the cells were detected by flow cytometry.Ratio of fluorescence intensity between peptide group and blank control group was used as an index to measure the level of HLA molecular remodeling,and then represented the interactions between peptide and HLA-B*1301 molecule.Results:HLA-polypeptide complex was denatalized and dissociated after being treated with pH3.0 buffer for 1 min,and only HLA heavy chain molecules were retained on the cell surface.The addition of peptides with binding force could significantly improve the level of HLA molecular remodeling,and the binding force of peptides with HLA-B*1301 could be evaluated by this method.Conclusion:HLA molecular reconstitution assay is a simple and reliable method to detect the binding of peptides to HLA-B*1301,which can also provide reference for the study of other low frequency HLA molecular antigen pre-sentation patterns.
