Purification and characterization of protective antigen (PAG) against Leptospira interrogans

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Protective antigen (PAG), glycolipid nature substance, was extracted from Leptospira interrogans serovar lai strain 017 with chloroform-methanol-water (1:2:0.8, V/V) and was partially purified by silica gel column chromatography. In SDS-PAGE, the PAG was not detected by Coomassie brilliant blue staining, but was observed as single smear-like band by silver staining. The band corresponded to 24-30 Kd of standard protein. Since outer envelope (OE) fraction showed the same band, suggesting that the PAG is one of components of OE. Immunogenicity and protective activity of the PAG were compared with those of OE. the PAG was able to induce agglutinating antibody specifically, which is important for protection in leptospirosis, and did not induce any antibodies against cellular antigens of Leptospira except for agglutinogen in comparison with OE and whole cells. Furthermore, sera of mice immunized with the PAG exhibited opsonic activity against Leptospira by measurement of chemical luminescence derived from reactive oxygen. The PAG possessed protective activity against lethal infection of Leptospira in hamsters and seemed to be useful as component vaccine against leptospiral infection.