Validation of a species-optimized enzyme-linked immunosorbent assay for determination of serum concentrations of insulin in dogs

Abstract

Background: Measurement of canine serum insulin has relied on methods developed to measure human insulin. A species-optimized test for measurement of serum insulin in dogs is now commercially available.

Objective: The purpose of this study was to validate the canine ELISA for determination of serum insulin concentration in dogs.

Methods: Precision was determined by evaluating intra- and interassay coefficient of variation (CV), and accuracy was determined by dilution and spike recovery studies. A method comparison study with samples from 34 clinically healthy dogs and 73 dogs examined for various illnesses and disorders ("patients") was performed using the canine ELISA and an ELISA for human insulin. Biologic relevance of the canine assay was evaluated by measuring insulin in samples collected from 8 healthy dogs after administration of glucagon. A stability study was preformed with 6 samples stored at 20°C, 4-8°C, and -20°C.

Results: For the canine ELISA, intra- and interassay CVs were 4.3-7.8% and 4.4-7.7%, respectively. Mean recovery after dilution was 99% and recovery after spiking with porcine insulin was 116%. The canine and human ELISAs correlated well (r(2) =.94 for healthy dogs, r(2) =.88 for patient samples). After glucagon injection serum insulin concentrations increased significantly in 8 dogs. Insulin was stable for 30 days in 6 serum samples stored at -20°C and in most samples for 8 days at 4-8°C. Insulin was stable for <3 days at room temperature (20°C).

Conclusions: The new canine serum insulin ELISA had good precision and accuracy and correlated well with the previously used assay.