open access

https://hdl.handle.net/2115/57304

A monoclonal antibody-based sandwich enzyme-linked immunosorbent assay for detection of secreted alpha-defensin

英語

Inflammatory bowel disease

2013-12-15

Academic press inc elsevier science

Analytical biochemistry

443(2)

124-131

Paneth cells at the base of small intestinal crypts secrete alpha-defensins, which contribute to innate immunity and shape composition of enteric microbiota. Efforts to establish a relationship between secreted alpha-defensins and disease have been hampered by a lack of sensitive assays to quantify luminal alpha-defensins. Here we report on a highly sensitive sandwich enzyme-linked immunosorbent assay (ELISA) for the mouse Paneth cell alpha-defensin cryptdin-4 (Crp4) in varied sources, including luminal contents rinsed from stomach to distal colon and fecal pellets. One pair of monoclonal antibodies (mAbs), selected from 10 rat hybridomas secreting Crp4-specific mAbs, was optimized for Crp4 detection and specificity in the sandwich ELISA. In CD1 mice, luminal Crp4 levels increased gradually from 6.8 +/- 5.2 ng/ml in proximal small intestine to 54.3 +/- 10.3 ng/ml in distal small intestine, and the peptide was detected in colonic lumen and feces. Secreted Crp4 was reduced significantly in feces of IL10 null mice, a model of inflammatory bowel disease (IBD) when compared with wild-type controls. This Crp4 sandwich ELISA enables accurate determinations of luminal alpha-defensins as biomarkers of Paneth cell function and enteric integrity in diverse disease states such as IBD, infectious disease, graft versus host disease, and obesity in association with dysbiosis of the intestinal microbiota. (C) 2013 Elsevier Inc. All rights reserved.

学術雑誌論文

Accepted Manuscript

0003-2697