Evaluation of effective factors in the optimization of immunization to achieve antibodies against RhD antigen in culture medium

Abstract Background and Objectives Antibody against RhD antigen is important in blood transfusion and clinical medicine. Therefore, in this study, the effective factors in the optimization of in vitro immunization to achieve immunized lymphocytes that produce antibodies were evaluated to obtain antibodies against RhD antigen in the culture medium. Materials and Methods In an experimental study peripheral blood mononuclear cells were isolated from O-Negative whole blood bags using the Ficoll method. These cells were treated with L-leucyl-L-leucine methyl ester (LLME) and were exposed to soluble and particulate RhD antigen, interferon-gamma (IFN-γ), interleukin-4 (IL-4) and CpGODN in a culture plate at 37°C for one week. After this period of incubation, in order to evaluate the production of antibodies from lymphocytes on the supernatant obtained from the culture medium, ELISA assay was performed. Results The use of IFN-γ, IFN-γ + IL-4 and IFN-γ + CpGODN was identified as the most optimal factors for the production of antibodies in an in vitro immunization (by producing antibodies with concentrations of 75.851 ± 36.831, 82.195 ± 31.293, and 66.134 ± 24.814 ngrml   in 10 exposures). Conclusions  The use of immunization optimization agents, has an effective role in the production of antibodies against RhD antigen in the in vitro culture.