Extraction And Analysis Of The Immunigenicity Of Capsular Polysaccharide (CPS) Of Vibrio Vulnificus FJ03-X2

Vibrio vulnificus is a zootoxic pathogen which does great damage to aquatic animals.Capsular polysaccharide(CPS)is one virulence factor and antigen of Vibrio vulnificus,which plays great importance in infecting host.To research the immunogenicity of the CPS of Vibrio vulnificus,the CPS of Vibrio vulnificus FJ03-X2 was extracted and purified.The components,molecular weight,virulence and immunogenicity of the CPS were analyzed.The ultraviolet absorption peak of CPS samples was found at 200nm with uv-vis absorption spectrophotometer,which coincides essentially with the characteristics of polysaccharides absorption peak.There are no obvious protein and lipopolysaccharide (LPS)bands on silver-stained sodium dodecyl sulfate-polyacrylamide electrophoresis (SDS-PAGE).Agarose gel electrophoresis of the CPS sample showed that it contained a little of nucleic acid.These results indicated that pure CPS was effectively purified.Detection of the polysaccharides after gel electrophoresis was carried out by combined alcian blue and silver staining.The molecular weight of the CPS is 311～20 KDa.The CPS contained high and low molecular weight ingredients.The former could show disperse zone on gel only by alcian blue stain dyeing with high contents and it's relative molecular weight was among 311～43 KDa.The other showed on gel by combined alcian blue staining and silver staining augentation with low contents and it's relative molecular weight was among 43～20 KDa.With the CPS injecting into the peritoneal cavity of mice,using the bacterial agglutination,ELISA and Dot ELISA to analyzed the immunogenicity of the CPS. The results showed that the mouse anti-CPS serum could agglute with FJ03-X2,and the titer is 2^4～5;in ELISA the titer of the mouse anti-CPS serum was 1:12 800.This primarily demonstrated that the CPS had immunogenicity towards the mouse and it could induce the differential humoral immune response on the mouse. The examination about virulence showed that the CPS did not have directly virulence.The CPS was injected into the peritoneal cavity of European eels(Anguillia anguillia).The result of Dot ELISA showed that the CPS.could not induce the differential humoral immune response of European eels.The examination of the challenge showed that the CPS could not induce the protective immune response of European eels.