Preparation of anti-PD-L1 nanobodies fused with C3Fab and their effect on plasma half-life
10.3760/cma.j.cn121382-20231030-00109
- VernacularTitle:融合C3Fab抗PD-L1纳米抗体的制备及其对血浆半衰期的影响
- Author:
Zhanxiong WANG
1
;
Meng LEI
;
Yichen DENG
;
Chu LOU
;
Tianning YANG
;
Qianqian HU
;
Jiangwei LI
Author Information
1. 新疆大学生命科学与技术学院，乌鲁木齐 830046
- Keywords:
Programmed death receptor-1;
Nanobody;
Half-life;
IgG binding peptide C3Fab
- From:
International Journal of Biomedical Engineering
2024;47(1):53-59
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To prepare the anti-programmed death-ligand 1 (PD-L1) nanoantibody P3C8-C3Fab by ligating with C3Fab and to investigate its role in plasma half-life.Methods:The C3Fab peptide derived from protein G was molecularly fused with the nanobody P3C8 by DNA recombination technology. The nanoantibody P3C8-C3Fab was inducibly expressed and purified in the E. coli BL21 strain, and the binding of it to PD-L1 protein, mouse IgG, and PD-L1-expressing tumor cells was detected by enzyme-linked immunosorbent assay (ELISA). The residual P3C8-C3Fab was detected in mouse serum at different times using double-antibody sandwich ELISA to assess the prolongation of the plasma half-life of PD-L1 nanobodies by C3Fab. Results:The nanoantibody P3C8-C3Fab was successfully constructed, and it could efficiently express itself in soluble form in BL21. The purified NbP3C8-C3Fab protein was obtained with a mass fraction of about 90% at a yield of 7.18 mg/L. The affinity of P3C8-C3Fab for PD-L1 protein and mouse IgG gradually increased with increasing mass concentration and showed a concentration correlation. The binding of P3C8-C3Fab to lung cancer A549 cells showed a concentration correlation. The concentration standard curve of P3C8-C3Fab in mouse serum showed a typical S-shape with a concentration correlation. The plasma half-life of P3C8 was only 0.44 h, while the plasma half-life of P3C8-C3Fab was 21.27-fold higher, up to 9.36 h.Conclusions:The linkage of C3Fab to the nanobodies of P3C8 can significantly prolong the plasma half-life of P3C8, which is valuable for the improvement of in vivo nanobody effects.
