Anti-DEFA5 Monoclonal Antibody Clones 1A8 and 4F5 Immunoreactive Bioassay for Diagnosing Inflammatory Bowel Disease.

ABSTRACT

Background: Robust evidence suggests that the aberrant expression of α defensin 5 protein (DEFA5) in colon inflammatory bowel diseases (IBDs) underlies the distinct pathogenesis of Crohn's colitis, can be exploited as a reliable diagnostic biomarker to differential diagnosis of Crohn's colitis (CC) from Ulcerative colitis (UC) in otherwise indeterminate colitis (IC). We evaluated the specificity of the commercially available anti-DEFA5 antibodies and showed further validation of their appropriateness for a given application is required. Methods: We established two mouse monoclonal DEFA5 antibody clones 1A8 and 4F5 by immunizing the mice with purified recombinant protein and validated the specificity, selectivity and cross reactivity in recognizing the endogenous and recombinant DEFA5 protein, especially for Immunohistochemistry, Western blot, Immunoprecipitation, or enzyme-linked immunosorbent assay. Results: Clones 1A8 and 4F5 recognized effectively the endogenous DEFA5 in active human diverticulitis (DV), UC, CC or IC disease samples, including transiently transfected HEK293T cells expressing DEFA5 with high degree of specificity and minimal non-confounding cross reactivity. Conclusions: 1A8 and 4F5 clones are worth studying in larger IBD cohorts to fully address whether DEFA5 expression may be used as a diagnostic biomarker to discrimination of the diagnosis of UC from CC or IC into authentic CC or UC or a colitis with different pathological characteristics.