[Development of a novel reporter gene method for determination of ADCC potency of anti-CD20 monoclonal antibody].
The biological activity of ADCC by anti-CD20 monoclonal antibody was determined by BioGlo™ Luciferase Assay System using Jurkat/NFAT-luc+FcγRIIIa cell line as effector cell and WIL2-S cell line as target cell. The developed method was verified for specificity, precision and accuracy. Anti-CD20 monoclonal antibody showed a dose-response mode by the developed method, and the determination result complied with the following four-parameter equation: y = (A-D)/[1 + (X/C)(B)] + D. The optimized parameters of the method were determined including the antibodies diluted concentration (18,000 ng·mL(-1)), dilution rate (1:5), the ratio of effector cell and target cell (6:1), and induction time (6 h). The values of eight independent tests have passed a statistical test for curve regression analysis, linear or parallelism, which showed the method possessed good specificity. Four different dilute groups of recovery rates sample were determined for 3 times, and the result showed mean relative potencies of (44.39±3.93)%, (72.74±2.78)%, (128.28±7.01)% and (168.19±2.70)% respectively, with a variation coefficient of less than 10%, and the recoveries of (88.78±7.85)%, (96.99±3.70)%, (102.63±5.61)% and (112.12±1.80)% respectively. A novel reporter gene method for determination of biological activity of ADCC by anti-CD20 monoclonal antibody was successfully developed, which showed strong specificity, good reproducibility and high accuracy, and might be used routinely.
Chun-yu
CY
Liu
Lan
L
Wang
Wei
W
Guo
Chuan-fei
CF
Yu
Feng
F
Zhang
Wen-bo
WB
Wang
Meng
M
Li
Kai
K
Gao
D058846:Antibodies, Monoclonal, Murine-Derived / Q000494:pharmacology*; D000920:Antibody-Dependent Cell Cytotoxicity; D018951:Antigens, CD20 / Q000276:immunology*; D017930:Genes, Reporter; D006801:Humans; D015203:Reproducibility of Results; D000069283:Rituximab
D058846:Antibodies, Monoclonal, Murine-Derived; D018951:Antigens, CD20; D000069283:Rituximab