Objectives: Chicken eggs present an ideal alternative antibody source to mammals, as the IgY in the chicken’s blood is transported to the egg and accumulates in the egg yolk in large quantities. The IgY have been recognized as an excellent source of polyclonal antibodies for over a decade. This simple non invasive approach presents an appealing alternative to conventional polyclonal antibody production methods. Therefore IgY is a promising candidate, both as an alternative to antibiotics and as a useful tool in research and diagnostics. In his study we aimed at the production, purification and characterization of egg yolk immunoglobulin (IgY) against antibiotic-resistant strains of Pseudomonas aerogenosa in laying hens. Materials & Methods: Samples were obtained from Ghaem Hospital, Mashhad, Iran. For confirmation of samples microbiological tests were performed .To determine the virulence of bacteria ,suspension of three concentrations of bacteria (1.5×106 ,1.5×107 ,1.5×108 CFU/ml) were injected intraperitoneally in mice. For IgY production, we used 3 hens in test and one in control group (HY-Line W-36 ,16-Week). Test group was immunized with 0.5 ml formalin killed P. aeruginosa vaccine(1.5×107 CFU/ml ) and for control group 0.5 ml sterile PBS mixed with 0.5 ml complete Freunds adjuvant was used. For the next two booster injections Incomplete Freund\'s Adjuvant was used. IgY was separated from egg by PEG6000 and characterized by dot-blot and ELISA methods. Results and Conclusions: Microbiological results including hemolysis around the colonies, colony odor, motility, hydrolysis of gelatin all confirmed the presence of inoculated bacteria. Intraperitoneally injected bacterial suspension showed that the lethal dose of bacteria was CFU 1.5×107. The results of SDS-PAGE electrophoresis showed protein bands of 27 and 67 kDa, which confirmed the presence of light and heavy chains of IgY. To determine the specificity of the antibodies produced in this study, dot-blot was performed which confirmed these findings. In the final step we used Indirect ELISA to determine the quantity of antibodies that bind specifically to the antigen with the concentration of 1/1000 p.aerogenosa antigen (50µl per well). After measurement of OD by ELISA reader, differences between test and control group data was analyzed by SPSS software Version 16 by Mann-Whitney U Test Statistics. Results show that specific antibodies were able to detect antigen in concentrations of 1 mg / ml, 0.5 mg/ml and 0.25 mg/ ml (P < 0.001). These findings indicate that eggs from hens immunized with appropriate antigens have potential as a useful source of passive immunity or for diagnosis and treatment of antibiotic resistant strains of P.aerogenosa at early stage of infections.

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