Heterogeneity Of Monoclonal Antibodies Research Articles

Heterogeneity of monoclonal antibodies against pancreatic beta cells.

Sixteen murine hybridoma-secreting monoclonal antibodies against pancreatic islet cell surface antigens (mc-ICSA) have been produced by the cell fusion technique using splenocytes from xenogeneic islet cell- or RIN cell-immunized Balb/c mice. In addition, some mice were autoimmunized by subdiabetogenic doses of the beta cell toxin streptozotocin in combination with complete Freund's adjuvant. Isotyping of the mc-ICSA revealed that 13 of the antibodies belong to the class IgM, and 3 to the subclass IgG1. The specificity of these mc-ICSA has been detected by means of the indirect immunofluorescent technique using primary rat islet cells suspensions, following a procedure of double immunostaining for pancreatic insulin and glucagon. Furthermore, the cross reactivity of these mc-ICSA was studied using endothelial, neuroblastoma and fibroblast cell lines and primary rat splenocytes. One out of the 10 mc-ICSA tested was alpha cell-specific, 2 were beta cell-specific and 7 out of 10 were reactive with both alpha and beta cells. Eleven mc-ICSA showed no cross-reactivity with the 5 other cell types tested. The binding of one mc-ICSA was blocked by 6 of the ICSA-positive human sera which were tested, suggesting that this monoclonal recognizes the same antigenic determinant. The same mc-ICSA diminished the glucose- and arginine-stimulated insulin secretion of isolated rat islets.

Characterization and Heterogeneity of Monoclonal Antibodies Directed to Intestinal Mucin Derived from Crohn's Disease Small Bowel

A panel of 12 monoclonal antibodies were produced by immunization of Balb/c mice with small bowel epithelial cells obtained from a patient with active well-documented Crohn's disease. The clones were derived by screening with immunofluorescence microscopy; those with staining patterns suggestive of mucin directed epitopes were chosen for study. Several distinct patterns of staining reactivity were noted, including reagents with homogeneous, luminal, heterogeneous and peripheral goblet cell activity. In addition, SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting analysis revealed reactivity to high molecular weight mucin. The reactive antigen was resistant to proteinase digestion. No endoneuraminidase activity was detected; however, one neuraminidase sensitive sialic acid epitope was demonstrated. Confirmation of glycoprotein epitopes was accomplished by testing purified mucins from several areas of the gastrointestinal tract by ELISA. Finally, individual small bowel goblet cell heterogeneity was demonstrated by immunofluorescence, Western blotting, and antibody affinity chromatography. These data demonstrate both by morphology and specific binding of antibody affinity chromatography a significant degree of small bowel goblet cell mucin heterogeneity.