In our report of a disease-specific colonic tissue-bound antibody (CCA) from patients with
ulcerative colitis (Proc Natl Acad Sci USA 1978;75:4528), crude CCA was largely fragmented, and the yield was small. We have modified the extraction procedure to increase the yield of intact
IgG present in CCA by sequential elution, storage of tissues in presence of a
protease inhibitor, 2 mM
Phenylmethylsulfonyl Fluoride, and use of
Phenylmethylsulfonyl Fluoride in extraction
buffers. Intact CCA-
IgG was purified using
protein A-Sepharose 4B affinity chromatography. 125I-CCA-IgG formed
immune complexes in vitro with the aqueous extracts of colonic mucosa from 5 patients with
ulcerative colitis but not from 6 patients with
Crohn's disease and 6 normal colons from patients with
carcinoma (p less than 0.01). We also performed reverse experiments by iodination of the colonic mucosal extracts with
Bolton-Hunter reagent and incubating them with several preparations of CCA-
IgG. Mucosal extracts of colon from 3 patients with
ulcerative colitis bound significantly with CCA-
IgG when compared with the identical extracts from 3 patients with
Crohn's disease, 3 with a normal colon (p less than 0.005), and with the control human
IgG (p less than 0.025). These studies demonstrate a better method of extraction and purification of intact CCA-
IgG. Intact CCA-
IgG binds to a specific
protein(s) present in the homogenates of colonic mucosa from patients with
ulcerative colitis but not from patients with
Crohn's disease and normal colon.