Development and Clinical Application of Indirect ELISA for Human Anti-Mouse Antibody in Human Serum

Objective To develop an indirect ELISA for human anti-mouse antibody(HAMA)in clinical serum specimens.Methods An indirect ELISA for HAMA in sera was developed,and the experimental condition was optimized,based on which the cutoff values for judgment of positive and negative results were determined,and the developed ELISA was verified methodologically.The serum specimens of healthy volunteers injected with McAb WuTac in clinical trial phase Ⅰwere determined by the developed ELISA.Results The optimal antigen coating concentration of the developed ELISA was 0.313 μg / ml,and the optimal dilutions of serum and enzyme-labeled antibody were 1 ∶ 160 and 1 ∶ 80 000 respectively.The cutoff A value of positive result was 0.210.The inter-and intra-CV values of the developed ELISA for determination of HAMA-positive specimens were 7.46% and 5.48%,and those for HAMA-negative specimens were 8.03% and 14.7%,respectively.The positive serum specimens confirmed by the developed ELISA showed no cross reaction with horse,goat,rabbit and monkey sera.HAMAs were induced in healthy volunteers 10~14 d after injection with WuTac.Conclusion An indirect ELISA for HAMA in human serum was successfully developed,which met the re-quirements for clinical trial of biological products in China.