| Abstract: | A regimen for the production of a monoclonal antibody that can distinguish between adult-expressed and fetal-expressed DNA polymerase a has been developed, but has not yet been completed. Hybrid BALB/c x C57BL/6 mice were immunized 3 x intraperitoneally with polymerase over a 2 month interval. During the first immunization, ace-mannan was also introduced as an immunomodulator to boost antibody production and secretion by plasma cells. Two weeks after the last injection, the spleen cells were fused separately with two non-immunoglobulin secreting murine myeloma cell lines, Sp2/0-Ag14 and P3X63/Ag8. 653. Four weeks later, the supernatant from two wells with hybridoma colonies were tested for antigen specificity by enzyme-linked immunosorbent assay (ELISA). Due to a lack of readily available antigen and an inaccurate standard antibody titration curve in the ELISA checkerboard titration, the cells were frozen at -70°C so that a more reliable ELISA system could be established without permitting cell overgrowth. Once ELISA systems for the two isozymes of DNA polymerase a are operational, the plates of hybridomas will be thawed, tested and cloned by limiting dilution until a monoclonal antibody is developed that can distinguish between the fetal and adult isozymes. |
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