PSI-15 Validation of a rapid test to evaluate humoral immunity in Labrador Retrievers across multiple life stages

Abstract This study sought to validate the D2Dx rapid immunity test in canines and investigate the age-related changes in the humoral immune system of healthy large breed dogs. Blood plasma with EDTA anticoagulant was collected from healthy Labrador Retrievers (n = 80) housed at Four Rivers Kennel under IACUC protocol FRK-56. Age groups were defined as follows: puppy 0-1.5 yr, adults 1.5-7 yr, senior 7-10, and geriatric 10-14. Immunity biomarkers including c-reactive protein (CRP), interleukin 4 (IL-4), interferon gamma (IFN-γ), complement protein 3 (C3), and immunoglobulin G (IgG) were determined through canine specific enzyme-linked immunosorbent assay (ELISA) kits. The nanoparticle based D2Dx test, which evaluates the cumulative humoral immune response, was also performed. Data were analyzed using a one-way ANOVA between age groups, with a Tukey’s post hoc, and correlations between variables were evaluated through multivariate analysis. The D2Dx score was correlated with body weight, age, IgG, and C3 validating that the test was able to adequately measure the humoral immunity in canines (r ≥ 0.49). D2Dx score was significantly different between age groups (P < 0.01) with puppies having significantly reduced scores compared with all other groups (P < 0.01). C3 was significantly different between age groups (P < 0.01), with puppies significantly less than other life stages (P ≤ 0.03). The C3 of adults averaged significantly greater than seniors and trended greater than geriatric subjects (P ≤ 0.01, 0.07). IgG between ages was also significantly different, with puppies significantly less than other age groups (P < 0.01). There was no significant difference between age groups for IFN-γ, IL-4 ratio, IFN-γ, IL-4, or CRP (P ≥ 0.18). This study showed that the D2Dx Rapid Immunity Test is able to detect changes to humoral immunity in canines. Beyond evaluating the effectiveness of the D2Dx test, this study also provides important information about the under researched topic of canine humoral immunity development and senescence in a robust, healthy cohort of large breed dogs.