Development of ELISA Method for Recombinant Hepatitis E Virus Antigen

Objective To develop a double antibody sandwich ELISA method for quantitative detection of recombinant hepatitis E virus(HEV) antigen.Methods Hens were immunized with virus-like particles(VLPs) of recombinant HEV,based on which a double antibody sandwich ELISA method was developed using the prepared polyclonal antibody against HEV-IgY as coating antibody and HRP-labeled monoclonal antibody against HEV as detection antibody,and verified.The HEV antigen contents in three batches of final products of HE vaccine were determined by the developed ELISA method,and the adsorption rate of aluminium hydroxide with HEV antigen was calculated.Results The linear detection range and minimum detection limit of developed ELISA method were 2 ～ 128 and 2 ng / ml respectively.The prepared ELISA kit showed no cross reactions with freeze-dried live attenuated hepatitis A vaccine,recombinant human IL-2,HBsAg,human serum albumin,calf serum or healthy human serum.The internal references for HEV antigen,at three concentrations,were detected by the developed ELISA method,and the results showed that the recovery rates were 95.13% ～ 104.50%,while both the intra-and inter-coefficients were less than 15%.No significant change was observed in the A450 value or sensitivity of internal reference for HEV antigen detected by using the HEV-IgY coated ELISA plate after storage at 37℃ for 5 d.All the adsorption rates of aluminium hydroxide with HEV antigen in three batches of final products of HE vaccine were more than 95%,which met the relevant requirements for quality control.Conclusion A double antibody sandwich ELISA method for quantitative detection of HEV antigen was successfully developed.