Preparation of Monoclonal Antibody and Development of Quantitative ELISA for Pertussis Filamentous Hemagglutinin

Objective To prepare monoclonal antibody(McAb) and develop an ELISA method for quantitative determination of pertussis filamentous hemagglutinin(FHA).Methods The McAbs against FHA was prepared by hybridoma technique and identified.The reaction condition of ELISA system was optimized,based on which double antibody sandwich ELISA for quantitative determination of FHA was developed and verified for precision and accuracy.The FHA contents in 12 batches of intermediate products of acellular pertussis vaccine were determined by the developed ELISA method.Results Four hybridoma cell strains stably secreting McAb against FHA were obtained,and the secreted McAbs were of IgG1 and IgG2b subtypes,with an ELISA titer of 1 ∶ 105 ～ 1 ∶ 106 in ascites,and showed specific reaction with FHA,while showed no cross reaction with pertussis toxin or influenza virus hemagglutinin.The linear detection range of developed ELISA method was 1.56 ～ 100.00 ng / ml,while both the intra-and inter-coefficients of variation were less than 10%.The recovery rates of FHA at high,moderate and low concentrations were 94.73%,108.67% and 116.37% respectively.The FHA contents(X ± 2SD) in 12 batches of intermediate products of acellular pertussis vaccine were stable.Conclusion The McAbs against FHA of pertussis were successfully prepared,and a double antibody sandwich ELISA method with high precision and accuracy was developed,which might be used for the determination of FHA content in intermediate products of acellular pertussis vaccine.