Immunoassays of human trefoil factors 1 and 2: measured on serum from patients with inflammatory bowel disease

Abstract

The trefoil factors (TFF1-3) are cysteine-rich peptides expressed in the gastrointestinal tract where they play a critical role in mucosal protection and repair. The expression is up-regulated at sites of ulceration in various chronic inflammatory diseases. Recently, we presented an ELISA method for measurement of TFF3. The aims of the present study were to develop and evaluate ELISAs for the other two known human trefoil peptides, TFF1 and TFF2, and to carry out a cross-sectional study on serum TFF levels in patients with inflammatory bowel disease (IBD). The TFF1-ELISA was based on two polyclonal rabbit antibodies and the TFF2-ELISA on a monoclonal mouse antibody and a polyclonal rabbit antibody. RhTFF1 and 2 were employed to prepare the calibrators. TFF1-3 were assayed in serum from IBD patients (n=41) and controls (n=13). The TFF1- (TFF2-) ELISA had a detection limit of 3 pmol/L (6 pmol/L) and an analytical imprecision (CV(A)) of 7.0-8.8 for mean concentrations of 24-120 pmol/L (6.1-8.0 for mean concentrations of 17-77 pmol/L). The central reference intervals (n=300) were 140-1400 pmol/L (37-190 pmol/L). There was no variation with age and menstrual cycle. Food intake reduced concentrations of TFF1 by approximately 15%, but did not influence concentrations of TFF2. TFF1 and TFF3 were increased in serum from IBD patients. We have developed assays for measuring TFF1 and TFF2. Finding increased TFF concentrations in serum from IBD patients suggests that measurements of trefoil peptides may be of clinical relevance in IBD.