Immunological effect of conjugated alginate-exotoxin A Pseudomonas aeruginosa on the titer of G antibody subclasses in Mice

Background and Aim : Pseudomonas aeruginosa is an opportunistic pathogen and alginate is its most important factor in pathogenicity. The objective of study is the immunogenicity evaluation of P. Pseudomonas aeruginosa alginate conjugated to exotoxin A as a vaccine candidate in mice Methods : mucoid strain of Pseudomonas aeruginosa ۶۴۹۴ was used to prepare alginate and the separation of exotoxin A was done with the standard strain of PAO۱. Alginate was extracted by the means of sedimentation with cold ethanol, dialysis, enzymatic digestion and chromatography. To improve immunogenicity, purified antigen was coupled to exotoxin A with ADH as a spacer and EDAC as a linker. Based on confirmation tests, resulting conjugate was devoid of specific toxicity and pyrogenic effect. Four groups of BALB/c female mice (each group was included ۱۵ mice) were selected in the next step. The first group with the ALG, the second group with the D-ALG-ETA and the third one with the ETA were vaccinated. The fourth group as a control group was vaccinated with normal saline. Vaccination was performed in three injectable doses with two-week intervals. Subsequently, serum samples were collected and antibody responses were measured by the ELISA method, for total IgG, IgG۱, IgG۲a, IgG۲b, IgG۳ Results : After the second and third doses with ALG-ETA showed a significant increase in antibody titer against ALG-ETA in comparison with pure ALG. The titers of IgG۲a, IgG۲b, IgG, IgG۱, IgG۳ antibodies that produced against alginate increased in the third injection compared to the first conjugate injection which was ۲/۹, ۹/۲, ۱۰, ۵/۲ and ۳ respectively.Conclusion : These results show that ALG from Pseudomonas aeruginosa increases antialginate antibodies in conjugate form with exotoxin A and can be considered an appropriate effective adjuvant.

کلیدواژه ها:

Pseudomonas aeruginosa ، Alginate ، Exotoxin A ، conjugate ، ELISA