Preparation of Monoclonal Antibodies With Hybridoma Techniques Against Promastigote of Leishmania infantum Antigens in Diagnosis of Visceral Leishmaniasis

Abstract

Background: Since the discovery of hybridoma cells, the uses of monoclonal antibodies (mAbs) are in vogue. Such antibodies with single isotype have high specificity. The developments in the field of cell culture and technology have led to the production of improved qualities of mAbs. In general, mAbs are important reagents used in biomedical research, as well as in targeted drug delivery systems. Objective: The aim of this study was to apply different strategies to produce mAbs against proamastigote Leishmania infantum strain in Iran. Materials and Methods: At first, standard strains were cultured and antigens of L. infantum were obtained. Afterward, BALB/c mice were immunized and antibody titers were determined. For hybridoma cell formation, isolated lymphocyte cells from spleen of immunized mice and myeloma cells were fused at the ratio of 10:1 in the presence of polyethylene glycol and followed by limiting dilution method for the isolation of monoclones. Results: More than 20 positive monoclones were hybridoma, from which 3 clones had optical density over 1.5. We named these clones as 5D2 FVI6, 3G2 FV7, and 3G2 FV5 which were selected for limiting dilution. From these hybrids, anti-promastigotes L. infantum mAbs were obtained. The results of isotype determination showed IgG2b sub-class (and not IgG1, IgG2a and IgA) in 5D2 FV and 3G2 FVI monoclones. Conclusion: This study produced mAbs against promastigotes of Iranian strain of L. infantum for the first time. These antibodies have reactivity against Iranian strain of promastigotes L. infantum and can be used in the diagnosis of visceral leishmaniasis.