Characterization of monoclonal antibodies to rabbit renal cortical cells

• Published in: The American journal of physiology Vol. 250; no. 3 Pt 1; p. C506
• Main Authors: Ronco, P, Geniteau, M, Poujeol, P, Melcion, C, Verroust, P, Vandewalle, A
• Format: Journal Article
• Language: English
• Published: United States 01.03.1986
• Subjects: Animals; Antibodies, Monoclonal - immunology; Antibodies, Monoclonal - isolation & purification; Antibody Specificity; Cell Fractionation; Electrophoresis, Polyacrylamide Gel; Epitopes; Female; Fluorescent Antibody Technique; Kidney Cortex - cytology; Kidney Cortex - immunology; Kidney Tubules - immunology; Microvilli - immunology; Rabbits
• ISSN: 0002-9513
• DOI: 10.1152/ajpcell.1986.250.3.c506

The immunological heterogeneity of the rabbit nephron was investigated using monoclonal antibodies. Seventeen antibodies have been produced by fusion of NS1 myeloma cells with spleen cells from BALB/c mice immunized with unfractionated rabbit renal cortical cell preparations. Sixteen antibodies reacted with proximal tubular cells: 11 with the brush border and 5 with basolateral membrane or intracytoplasmic components. Only one of the latter was specific for constituents of the proximal tubule. One antibody reacted with the cortical collecting tubule. Eight of the anti-brush-border antibodies were further characterized by immunoprecipitation of detergent-solubilized radiolabeled brush-border membrane vesicles. Seven proteins with subunits ranging in molecular weight from 90,000 to greater than 340,000 were identified. Systematic survey showed that one of these proteins with a subunit molecular weight of 115,000 exhibited leucine aminopeptidase activity. Selected monoclonal antibodies bound to Sepharose 4B immunoadsorbents were used to deplete solubilized brush-border membrane vesicles of a given antigen and to identify leucine aminopeptidase. Furthermore, the obtention of specific antibodies directed against the proximal tubule allowed us to set up a simple method for renal cell separation: isolated renal cortical cells could be depleted by 80% in proximal cells by passage over columns of Sepharose 6MB covalently linked with three different monoclonal anti-brush-border antibodies, thus leading to cell suspensions considerably enriched in tubule cells originating from the more distal segments of the nephron.