Preparation of human monoclonal antibody against SARS-CoV-2 spike protein using single B cell
- VernacularTitle:利用单个B细胞制备人源SARS-CoV-2刺突蛋白单克隆抗体
- Author:
FENG Ze⁃zhong
1
;
LU Yang
;
LI Jia⁃ying
;
MA Ping
;
WANG Ying⁃nan
;
ZHU Jin⁃qi
;
SUN Jin⁃fu
Author Information
- Publication Type:Journal Article
- Keywords: Severe acute respiratory syndrome coronavirus 2（SARS⁃CoV⁃2）；Single B cell；Monoclonal antibody；S protein
- From: Chinese Journal of Biologicals 2023;36(1):48-52
- CountryChina
- Language:Chinese
- Abstract: Abstract：Objective To prepare human monoclonal antibody against spike protein（S protein）of severe acute respiratory syndrome coronavirus 2（SARS⁃CoV⁃2）by using single B cell，and determine its neutralizing activity. Methods Venous blood with high antibody level was collected from people immunized with inactivated SARS⁃CoV⁃2 vaccine（Vero cells） twice，of which peripheral blood mononuclear cells（PBMCs）were isolated by lymphocyte stratified fluid and used to isolate single B cell expressing S protein antibody by magnetic beads coupled with S1 protein. Variable region genes of IgG heavy chain and light chain were amplified by nested PCR after reverse transcription of single B cell，which were connected with CMV promoter，IgG leader sequence，IgG constant region and polyA sequence by overlapping PCR to construct antibody linear expression cassette. Linear expression cassette of the heavy chain and light chain from the same B cell was transfected to HEK293T cells to express human monoclonal antibody of SARS⁃CoV⁃2 S protein. Immunoreactivity was detected by immuno⁃ fluorescence while neutralizing activity by pseudovirus neutralization test. Results A total of 26 monoclonal antibodies against SARS⁃CoV⁃2 S protein were expressed，which showed heavy chain and light chain protein bands of IgG antibody at
- Full text:利用单个B细胞制备人源SARS-CoV-2刺突蛋白单克隆抗体.pdf
