Development and verification of a double antibody sandwich ELISA method for determination of relative potency in vitro of inactivated SARS-CoV-2 vaccine(Vero cells)
Objective To develop and verify a double antibody sandwich ELISA method for the determination of relative potency in vitro of inactivatedSARS-CoV-2 vaccine(Vero cells).Methods Using cell fusion technology to prepare SARS-CoV-2 monoclonal antibodies,the sandwich ELISA method with the monoclonal antibodies as the coating antibody and SARS-CoV-2 polyclonal antibodies as the detection antibody was developed to detect the in vitro relative potency of inactivated SARS-CoV-2 vaccine(Vero cells),and the linear range,specificity,precision,and durability of the method were verified.The in vitro relative potency of 20 batches of inactivated SARS-CoV-2 vaccine(Vero cells)was detected using the developed method,and the correlation with the corresponding in vivo potency results was analyzed.Results The monoclonal antibody 20D8 was selected for the development of the method,with a purified titer of up to 105 and a protein concentration of 2.69 mg/mL.The reference of inactivated SARS-CoV-2 vaccine(Vero cells)showed a good linear relationship with A450/630 at the concentration of 3.2to200WU/mL.The linear equation was y=0.846 8 x-1.483,with an R2 value of 0.991 9.The developed method specifically detected inactivated SARS-CoV-2 vaccine(Vero cells)without cross-reactivity with other vac-cines.The CVs of repeatability verification and intermediate precision verification were 9.91％and 11.03％,respectively.The in vitro relative efficacy CV of inactivated SARS-CoV-2 vaccine(Vero cells)sample was 8.59％after post-dissociation at 37 ℃ for 22,24 and 26 h,and 6.62％after post-dissociation at 36,37 and 38 ℃ for 24 h.The relative potency in vitro of 20 batches of inactivated SARS-CoV-2 vaccine(Vero cells)was positively correlated with the results of potency in vivo(r=0.7,P＜0.05).Conclusion The developed sandwich ELISA method has good specificity,precision and tolerance,and is easy to operate,which can be used for the detection of the in vitro relative efficacy and the quality control of inactivated SARS-CoV-2 vaccine(Vero cells).
NETL
NSTL
万方数据
