Preparation of antibody against alpha-toxin in guinea pigs and detection of yeast display vaccine / 中国医师进修杂志

ABSTRACT

Objective@#To prepare anti-serum against α-toxin in guinea pigs and purify anti-rHla IgG and identify it by SDS-PAGE observe the expression of Hla on the surface of yeast cells by ELISA and immunofluorescence confocal microscopy.@*Methods@#Guinea pigs were immunized with recombinant α-toxins to obtain anti-α-toxin serum and total IgG which contains anti-rHla IgG. The purity of IgG was Identified and evaluated with non-reducing SDS-PAGE. The expression of Hla was evaluated with whole cell ELISA and immunofluorescence confocal.@*Results@#The anti-α-toxin antibody of guinea pig was successfully obtained. Anti-α-toxin antibodies was captured by the protein A on the pre-packed column. The concentration of IgG in the unpurified serum was low, the α-rHla content in the flow-out peak was even lower, and the elution fraction contains IgG (α-rHla) with purity of about 85%. Through ELISA, transformant-1 and transformant-2 was identified to be positive with rHla on the surface, compared with that in the negative control. After stain with the purified anti-α-toxin IgG, the 1 yeast transformant showed green fluorescence under immunofluorescence confocal microscope. By contrast, without inducing, the 1 yeast transformant was negative with fluorescence.@*Conclusions@#The results show that Staphylococcus aureus Hla is successfully displayed on the surface of yeast, and this research could be further applied in the identification of clinical samples or pre-clinical research. This study provides two reliable methods for the development of yeast-display vaccine.