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Development and validation of an enzyme-linked immunosorbent assay for feline trypsin-like immunoreactivity
J M Steiner1, D A Williams, E M Moeller
1Department of Small Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station 77843-4474, USA.
Objective:
To develop and validate an ELISA for quantitative analysis of feline trypsin-like immunore-activity (fTLI).
Sample Population:
Purified feline cationic trypsin (fCT) and rabbit anti-fCT antiserum; blood samples from 63 healthy cats.
Procedures:
A sandwich capture ELISA was developed, using anti-fCT antiserum purified by affinity chromatography that underwent biotinylation. Purified fCT was used for standards. The assay was validated by determination of sensitivity, working range, linearity, accuracy, precision, and reproducibility. A reference range was established by assaying serum samples from the 63 healthy cats.
Results:
Sensitivity was 1.23 microg/L; working range was 2 to 567 microg/L. Ratios of observed versus expected results for 4 samples tested at various dilutions ranged from 90.0 to 120.7%. Ratios of observed versus expected results for 5 samples spiked with various concentrations of fCT ranged from 82.0 to 101.8%. Intra- and inter-assay coefficients of variability ranged from 9.9 to 11.1% and from 10.2 to 21.7%, respectively. The reference range for serum fTLI measured with this ELISA was 12 to 82 microg/L.
Conclusions And Clinical Relevance:
Results suggest that an ELISA can be used to measure serum fTLI in cats. The ELISA was sufficiently sensitive, linear, accurate, precise, and reproducible for clinical use.