Distribution of O-acetylated sialomucin in the normal and diseased gastrointestinal tract shown by a new monoclonal antibody.
OBJECTIVE
To produce and characterise a monoclonal antibody specific for O-acetylated sialomucin and to assess its use in immunohistochemistry on a panel of normal and diseased intestinal tissue samples.


METHODS
Mouse monoclonal antibodies were developed following immunisation with highly purified human colonic mucin. One of these (MMM-17) showed strong binding to mucin throughout the normal colon with relative lack of binding to colon cancer tissue. The binding epitope of MMM-17 was then characterised by screening for agglutination activity against a panel of human and animal erythrocytes and by assessment of its binding to a range of normal and chemically treated slot blotted mucins. Further immunohistochemical studies were then performed on formalin fixed, normal, and diseased human intestinal samples.


RESULTS
Binding of MMM-17 to slot blotted human colonic mucin was reduced by 38 (SD 14%) (n = 4) by alkali treatment of the mucin, sequential alkali and sialidase treatment completely abolished binding. Sialidase treatment alone, however, caused only an 11 (11%) reduction in binding. MMM-17 failed to agglutinate any human, rabbit, rat or mouse erythrocytes. These findings were compatible with specificity of MMM-17 for sialomucins O-acetylated at the C-7 or C-8 positions on the sialic acid. Strong staining by MMM-17 was found in all goblet cells throughout all 40 normal colonic and rectal samples studied, but staining was absent in seven of 13 colorectal carcinomas. Normal duodenum (n = 16) and normal ileum (n = 3) all showed occasional positive goblet cells. The normal gastric antral mucosa was generally negative B MMM-17, but in all of 15 cases of gastritis with intestinal metaplasia the metaplastic glands were strongly positive for MMM-17.


CONCLUSIONS
Monoclonal antibody MMM-17 has specificity for O-acetylated sialomucins and its binding depends both on the position of O-acetylation and on the adjacent oligosaccharide structure. Preliminary studies using the antibody on archival tissue samples support the previous reports of reduced O-acetylation in colon cancer demonstrated by indirect histochemistry and show the neo-formation of O-acetylated sialomucin in intestinal metaplasia in the stomach.
Milton
J D
JD
Department of Medicine, University of Liverpool.
Eccleston
D
D
Parker
N
N
Raouf
A
A
Cubbin
C
C
Hoffman
J
J
Hart
C A
CA
Rhodes
J M
JM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D000918:Antibody Specificity; D003110:Colonic Neoplasms / Q000737:chemistry; D004064:Digestive System / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay; D005753:Gastric Mucosa / Q000473:pathology; D005767:Gastrointestinal Diseases / Q000378:metabolism*; D006801:Humans; D007124:Immunoenzyme Techniques; D008679:Metaplasia / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D009077:Mucins / Q000032:analysis*; D052116:Sialomucins
D000911:Antibodies, Monoclonal; D009077:Mucins; D052116:Sialomucins