In the present study, we examined the feasibility of using recombinant antibodies containing murine variable regions and human constant regions as calibrators or controls in immunoassays. As a model system, we chose the Abbott IMx Toxo immunoglobulin M (IgM) and Toxo IgG assays designed to detect antibodies to Toxoplasma gondii. Two mouse monoclonal antibodies were selected based on their reactivity to the T. gondii antigens P30 and P66. Heavy- and light-chain variable-region genes were cloned from both hybridomas and transferred into immunoglobulin expression vectors containing human kappa and IgG1 or IgM constant regions. The constructs were stably transfected into Sp2/0-Ag14 cells. In the IMx Toxo IgG assay, immunoreactivity of the anti-P30 chimeric IgG1 antibody paralleled that of the positive human plasma-derived assay calibrators. Signal generated with the anti-P66 chimeric IgG1 antibody was observed to plateau below the maximal reactivity observed for the assay calibrator. Examination of the IgM chimeric antibodies in the IMx Toxo IgM assay revealed that both the anti-P30 and anti-P66 antibodies matched the assay index calibrator manufactured with human Toxo IgM-positive plasma. When evaluated with patient samples, the correlation between results obtained with the chimeric antibody calibrators and the positive human plasma calibrators was ≥0.985. These data demonstrate that chimeric mouse-human antibodies are a viable alternative to high-titer positive human plasma for the manufacture of calibrators and controls for diagnostic assays.

The generation of stable rabbit-rabbit hybridomas is now possible by the recent development of a rabbit fusion partner. The ability to generate rabbit monoclonal antibodies (MAbs) can be advantageous because these rabbit immunoglobulins tend to exhibit higher affinity than murine MAbs. Furthermore, it has been observed that, in general, rabbits will elicit an immune response to antigens of limited immunogenicity in mice. Unfortunately, these rabbit-rabbit hybridomas secrete only 200 ng/mL to 5 microg/mL of immunoglobulin, which may limit larger scale production of rabbit antibodies. This study sought to determine if interleukin 6 (IL-6), which has been reported to have proliferative and secretory stimulating effects on some murine hybridomas, had any effect on a rabbit cell line that secretes a monoclonal IgG specific for estradiol. The results demonstrated that recombinant human IL-6 had a dose-dependent enhancing effect on the IgG secretion of the rabbit-rabbit hybridoma. The enhancing effect was consistent when the cells were continuously passed in the presence of IL-6. However, IL-6 did not affect the growth of the hybridoma. In contrast, no discernible effect was accomplished with recombinant mouse IL-6. Furthermore, no basal IL-6 activity was detected in the rabbit hybridoma extracellular medium. The IL-6 enhancement effect observed in this study may help to increase the immunoglobulin yield of rabbit hybridomas and to assist in the understanding of the mechanism(s) behind the lowered secretion level.

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