Monoclonal Antibody Production against Aspergillus Fumigatus in Ascetic Fluid of Balb/C Mice

Monoclonal antibodies (mAbs) due to their binding specificity, their homogeneity and great ability to be produced in unlimited quantities are important reagent in diagnosis and treatment of diseases like infections and cancers. Hybridoma cell lines grown as ascites tumours in Pristane primed mice will frequently yield milligram quantities of monoclonal antibody per milli liter of ascites fluid. For large scale production of monoclonal antibodies, hybridoma cells that produce monoclonal antibody against Aspergillus fumugatus were injected into the peritoneum of the Balb/c mice which have previously been primed with 0.5 mL Pristane. After 10 days, approximately 4-5 mL ascitic fluid was harvested from the peritoneum of each mouse. Ascitic fluid was assayed for the titer of monoclonal antibody in reaction with crude extract of Aspegillus fumigatus and its cross reactivity in reaction with other pathogenic species of Aspergillus and Candida. The titer of mAb was 100,000 and didn’t show cross reactivity with other pathogenic species of Aspergillus and Candida. Immunoblotting was used to study the specificity and nature of epitopes recognized by the antibodies and also, for confirmation of the result of cross reactivity with other species of Aspergillus and Candida by ELISA method. Periodate oxidation of crude extract of Aspergillus fumigatus on a PVDF membrane resulted in reduction of carbohydrate residue of the glycoproteins, and appearance of sharp bands. The proteins binding had a molecular mass of 58 kDa. The subclass of antibody was IgG1 and its light chain was kappa. Ascitic fluid was purified by ion exchange chromatography.