Anti-human neutrophil murine monoclonal antibodies have been selected which inhibit yeast induced respiratory burst. At least some are directed at the cell surface C3bi receptor and a group of related glycoproteins. A study is planned in which such monoclonal antibodies will be used as specific probes to study the structure, function, distribution and interaction of cellular components important for PMN activation. The subunit structure of the C3bi receptor in neutrophils will be examined by immunoprecipitation and two dimensional gel analysis. The functional effects of antibodies binding to different portions of the receptor complex will be compared. Changes in receptor structure and subcellular distribution during PMN activation will be investigated by cell fractionation and electron microscopy. The role of the receptor in triggering excitatory signals such as membrane depolarization and calcium flux will be studied. Interaction of the C3bi receptor with other functional cellular components including ligand receptors, the cytoskeleton, and the neutrophil NADPH oxidase will be assessed by fluorescence immunomicroscopy, nonionic detergent extraction, and immunoprecipitation experiments. The influence of the receptor on neutrophil-fungal interactions will be evaluated. Finally, attempts will be made to generate monoclonal antibodies directed at other functionally important cellular structures by using immunogens which are enriched for antigens of interest and very specific screening assays. It is hoped that this information will contribute to a better understanding of the polymorphonuclear leukocyte and its role in the inflammatory process.
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