Development and analytical validation of a radioimmunoassay for the measurement of alpha1-proteinase inhibitor concentrations in feces from healthy puppies and adult dogs.
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abstract
Canine (1)-proteinase inhibitor (c(1)-PI), a proteolysis-resistant protein with a molecular weight similar to albumin, has been shown to be clinically useful as a marker for gastrointestinal protein loss in dogs. A competitive, liquid-phase radioimmunoassay was developed and analytically validated. Fecal samples were collected from 101 healthy pet dogs of various breeds and ages, and fecal c(1)-PI (Fc(1)-PI) concentrations were compared between dogs of different age groups. A reference interval for Fc(1)-PI concentration was calculated using the central 95th percentile. Analytical sensitivity of the assay was 2.2 g Fc(1)-PI/g feces. Observed-to-expected ratios for the serial dilution and spiking recovery of 9 and 6 fecal extracts ranged from 90.4 to 152.0% and from 71.3 to 112.3%, respectively. Coefficients of variation for intra- and interassay variability for 6 fecal extracts were 10.8% and 12.5%, respectively. The reference intervals for the mean and maximum Fc(1)-PI from fecal samples collected on 3 consecutive days were 2.2-13.9 g/g and 2.2-21.0 g/g, respectively. Fc(1)-PI was significantly higher in dogs <1 year of age (P < 0.0001 for both mean and maximum Fc(1)-PI for the 3 samples). The radioimmunoassay described is sensitive, linear, precise, reproducible, and accurate for clinical use, thus allowing reliable quantification of Fc(1)-PI in clinical patients. Using this assay, a mean or a maximum Fc(1)-PI for 3 sampling days of >13.9 g/g or >21.0 g/g, respectively, should be considered abnormal in dogs >1 year of age. Fecal c(1)-PI concentrations in dogs <1 year of age were significantly higher and should be carefully interpreted in this age group.
