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Development and analytic validation of a radioimmunoassay for the quantification of canine calprotectin in serum and feces from dogs

Romy M. Heilmann Dr med vet1, Jan S. Suchodolski Dr med vet, PhD2, and Jörg M. Steiner Dr med vet, PhD3
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  • 1 Gastrointestinal Laboratory, Department of Small Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843-4474.
  • | 2 Gastrointestinal Laboratory, Department of Small Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843-4474.
  • | 3 Gastrointestinal Laboratory, Department of Small Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843-4474.

Abstract

Objective—To develop and analytically validate a radioimmunoassay (RIA) for the quantification of canine calprotectin (cCP) in serum and fecal extracts of dogs.

Sample Population—Serum samples (n = 50) and fecal samples (30) were obtained from healthy dogs of various breeds and ages.

Procedures—A competitive, liquid-phase, double-antibody RIA was developed and analytically validated by assessing analytic sensitivity, working range, linearity, accuracy, precision, and reproducibility. Reference intervals for serum and fecal cCP concentrations were determined.

Results—Sensitivity and upper limit of the working range were 29 and 12,774 μg/L for serum and 2.9 and 1,277.4 μg/g for fecal extracts, respectively. Observed-to-expected ratios for serial dilutions of 6 serum samples and 6 fecal extracts ranged from 95.3% to 138.2% and from 80.9% to 118.1%, respectively. Observed-to-expected ratios for spiking recovery for 6 serum samples and 6 fecal extracts ranged from 84.6% to 121.5% and from 80.3% to 132.1%, respectively. Coefficients of variation for intra-assay and interassay variability were < 3.9% and < 8.7% for 6 serum samples and < 8.5% and < 12.6% for 6 fecal extracts, respectively. Reference intervals were 92 to 1,121 μg of cCP/L for serum and < 2.9 to 137.5 μg of cCP/g for fecal extracts.

Conclusions and Clinical Relevance—The RIA described here was analytically sensitive, linear, accurate, precise, and reproducible for the quantification of cCP in serum and fecal extracts. This assay should facilitate research into the clinical use of serum and fecal cCP measurements in dogs with inflammatory bowel disease.

Contributor Notes

Dr. Heilmann's present address is Veterinary Referral Center TÜZ, Hauptstrasse 21, CH-4456 Tenniken, Switzerland.

The authors thank Niels Grützner for assistance with collection of samples and Kathleen M. Aicher and Dr. Nora Berghoff for technical assistance with the radiolabeling procedure.

Presented in part at the 2007 Annual Forum of the American College of Veterinary Internal Medicine, Seattle, June 2007.

Address correspondence to Dr. Steiner.