Development and Comparison of Methods for Determination of Biological Activity of Trastuzumab

Objective To develop and compare four methods for determination of biological activity of Trastuzumab.Methods BIAcore method for Trastuzumab was developed by surface plasmob resonance(SPR);a method for determination of activity of Trastuzumab in inhibiting the proliferation of human breast cancer SKBR3 cells was developed;a SKBR3 cells-based method for determination of binding activity of Trastuzumab as well as an ELISA method for determination of binding activity of Trastuzumab to ErbB2 antigen were developed.The four methods were verified for reproducibility and compared.Results The optimal pH value and sample concentration for BIAcore method were 5.0 and 0.05 ～ 5.00 nmol / L respectively,while the kinetic constant(KD) was 0.19 nmol / L.The median effective concentration(EC50) of Trastuzumab for inhibiting the proliferation of SKBR3 cells in vitro was 0.145 μg/ml(R2 0.98).The SKBR3 cells-based binding activity(EC50) of Trastuzumab was 0.124 μg/ml(R2 0.98).The binding activity(EC50) of Trastuzumab to ErbB2 antigen was 0.143 μg/ml(R2 0.98).All the four methods showed high reproducibility,by which the determination results were basically in agreement.Conclusion All the developed methods reflected the biological activities of Trastuzumab effectively,which provided several tools for rapid and simple screening of monoclonal antibodies.