Equine chorionic gonadotrophin isoform composition in commercial products compared to isoform composition in pregnant mare plasma

Abstract

It is well documented that there is considerable batch to batch variation in the activity of commercial preparations of gonadotrophins. These products are used in a variety of assisted reproductive procedures in the livestock industry, consequently this high degree of variation between products and batches adds to the already considerable between animal variations in response to the treatment. Equine chorionic gonadotrophin (eCG) is a heterodimeric glycoprotein hormone secreted by the placental endometrial cups during the first third of gestation in the horse. Plasma is harvested from pregnant mares between 40–90 days of gestation and the eCG isolated and used to formulate commercial preparations. Previous research has shown that eCG like the other gonadotrophins is a highly heterogeneous molecule with significant differences in bioactivity between isoforms. The aim of this study was to determine whether significant differences in isoform composition exist between various commercial preparations of eCG (n = 15), and how this compares with the isoform composition found in plasma (n = 23). Concentrations of eCG were determined using a competitive eCG ELISA. Liquid phase iso-electric focusing was used to fractionate plasma and the commercial preparations into 10 pH ranges from pH 3.0 to pH 10.0. Data from the 10 fractions were grouped into acidic (pH 3.0–5.1), intermediate (pH 5.2–7.9), or basic (pH 8.0–10.0) isoform categories for analysis. Immunoactivity between commercial eCG products ranged from 44% to 362% of stated bioactivity. Iso-electric focusing showed that the majority of the immunoactivity (92%) of the commercial preparations was found in the acidic fractions (pH 3.0–5.1), and in particular in the pH range 3.0–3.8. This contrasted starkly with isoform profiles found in pregnant mare plasma samples which showed a much greater spread across all 3 pH ranges. In summary, the isolation processes of commercial eCG preparations appears to selectively favour the acidic isoforms of eCG.