Reference intervals for equine fecal calprotectin and calprotectin:protein ratio and preliminary clinical evaluation in horses with colic and colitis

OBJECTIVE To establish the reference interval (RI) of fecal calprotectin (fCP) and fCP:protein ratio in the feces of healthy horses and demonstrate preliminary clinical utility for the quantification of intestinal inflammation. METHODS Feces were collected from healthy horses (n = 103) and horses with colic (n = 15) or colitis (n = 13). Feces were suspended in buffer to create fecal supernatant. Fecal calprotectin concentration was determined by ELISA, fecal total protein concentration was determined by bicinchoninic acid assay, and the fCP:protein ratio was calculated. Reference intervals for fCP and fCP:protein ratio were calculated from healthy horses. Descriptive and inferential statistics were performed to evaluate associations between fCP concentration and fCP:protein ratio with group and other clinical features. The study period was from September 20, 2023, through April 20, 2024. RESULTS The RI for fCP was 0.056 to 0.278 μg/mL; 4 of 13 colitis cases had fCP above the RI. The RI for fCP:protein was 6.6 X 10⁻⁶ to 4.9 X 10⁻⁵; 5 of 13 colitis cases and 2 of 15 colic cases had fCP:protein above the RI. Horses with colitis had significantly greater fCP (median [quartile 1, quartile 3], 0.234 [0.194, 0.279] μg/mL) than healthy horses (0.196 [0.138, 0.220] μg/mL) or colic cases (0.189 [0.164, 0.215] μg/mL). There was a greater difference in fCP:protein ratio between colitis cases (4.31 X 10⁻⁵ [3.19 X 10⁻⁵, 6.23 X 10⁻⁵]) and both healthy horses (2.19 X 10⁻⁵ [1.58 X 10⁻⁵, 2.88 X 10⁻⁵]) and colic cases (1.52 X 10⁻⁵ [1.22 X 10⁻⁵, 3.56 X 10⁻⁵]). CONCLUSIONS Both fCP and fCP:protein ratio were increased in horses in colitis; additional work is needed to establish clinical utility. CLINICAL RELEVANCE There are few reliable diagnostic modalities for inflammation of the equine large colon. With additional development of a clinically applicable test and validation in larger population, fecal calprotectin may facilitate noninvasive quantification of intestinal inflammation in horses.