Quantitative immunoassay for mink immunoglobulin in serum and milk

Abstract

Background: The significance of maternal immunoglobulin G (IgG) for the resistance against a number of infections affecting the health of young mink offspring is not known. Here, we present a validated immunoassay for quantification of mink IgG in serum and milk, using a commercially available polyclonal goat anti-ferret IgG antibody cross-reactive with mink IgG as both the catching and the detection antibody, in a sandwich format enzyme linked immunosorbent assay (ELISA). Using this ELISA, serum IgG concentrations was analyzed over time in both mothers and kits in order to establish a correlation between maternal IgG serum concentrations and those of the offspring. Results: Intra-assay coefficient of variation (CV) for a serum sample ranged from 2.15 to 5.97% depending on the dilution, while the inter-assay CV ranged from 5.17 to 17.78%. In addition, the range of milk intra-assay CV was 2.71-5.92%, while the range of the inter-assay CV was 4.20-16.03%. Calibrating the ELISA with purified mink IgG (an in-house preparation purified from mink serum) the lower limit of detection was found to be 5 ng/mL for serum and 1 ng/mL for milk. Both serum and milk showed high precision and good linearity over a two-log dilution range. When comparing the serum IgG concentrations of the mink kits a clear within litter effect was seen, while the mean serum IgG concentrations of litters differed significantly between some of the litters (P = 0.0013). Mean maternal serum IgG concentrations correlated positively with the IgG serum concentration of the corresponding offspring sampled over a 3 week period (R2 = 0.63). Conclusions: A calibrated and reproducible sandwich ELISA for quantifying mink IgG concentrations in both milk and serum with high analytical sensitivity was developed and validated. The results in this study corroborate previous investigations supporting the usability of the ELISA, paving the way for investigations into the importance of maternal IgG in milk and in serum for the welfare and health of the offspring.