The final optimal conditions for the PcAb based ELISA and the McAb based ELISA were as follows: the coating concentrations of R-VTG were 250 and 200 ng mL1, respectively, to decrease the consumption of this protein; the same dilution factor of primary antibody (1:64 000) was used; and the dilution factors of second antibody were 1:20 000 and 1:10 000, respectively.