Emergence of antigenic glycoprotein structures in ulcerative colitis detected through monoclonal antibodies

The emergence of new glycoprotein structures in colonic mucosa from patients with ulcerative colitis (UC) was assessed through the development of monoclonal antibodies (MAbs). Hybridomas were prepared from mice immunized with mucin glycoproteins purified from UC colonic tissue. Supernatants of 11 fusion products among 1200 fusion products that were screened in a solid-phase binding assay differentially bound UC-derived colonic mucin glycoproteins relative to comparable preparations from normal or Crohn's colitis tissue. These hybridomas were double-cloned to yield MAbs designated as MAbs UC 1-11. Disease-related specificity of MAbs UC 1-11 was determined through assessment of binding to beads coated with mucin glycoproteins purified from individual samples of UC tissue (n = 15), normal tissue (n = 21), and Crohn's colitis tissue (n = 10). Monoclonal antibody UC 7, an MAb of immunoglobulin G_2A subclass, showed differential binding in solid-phase assays to UC mucin glycoproteins, with a mean binding of 10,170 ± 2740 cpm per UC glycoprotein-coated bead versus 2300 ± 1080 and 2470 ± 1525 cpm for normal and Crohn's colitis-derived glycoproteins, respectively. Monoclonal antibody UC 11 showed similar differential binding to UC mucin glycoproteins (9860 ± 680 cpm vs. 1770 ± 420 cpm). Binding specificity in solid-phase assay was mirrored by colonic mucosal staining patterns assessed by indirect immunofluorescent staining. Monoclonal antibody UC 7 specifically stained colonie mucosa from 8 of 10 patients with active UC, none of the samples from 8 normal controls, and none of the samples from 11 disease controls (six with Crohn's colitis, five with other inflammatory disorders). Specific staining was present on both the epithelial surface and on cells scattered within the lamina propria. Staining by MAb UC 7 was also observed in 3 of 4 samples of proximal uninvolved mucosa from patients with left-sided ulcerative colitis and in 3 of 5 samples from UC patients without acute disease activity.