Development of MS-based methods for identification and quantification of proteins altered during early pregnancy in dogs

摘要

Increased knowledge on serum protein profiles during early pregnancy in dogs would be valuable for several reasons, including animal welfare. Inflammatory changes during this period have been described. Today, mass spectrometry (MS) is a well-established technique to perform unbiased qualitative and quantitative studies of proteins in body fluids regardless of species. In the present study, a shotgun proteomic analysis based on nano-liquid chromatography-MS was performed to identify proteins of altered abundance during canine pregnancy, and, thereafter, a targeted parallel reaction monitoring (PRM)-method was developed and applied to absolutely quantify the concentrations of a selection of these proteins. Among the 32 proteins found altered between pregnant and non-pregnant dogs in the initial analysis, 12 were selected based on their changes in concentration and known biological importance, and these were analyzed using the PRM method. The PRM method showed good linearity, repeatability and sensitivity, and confirmed the higher concentration of Fibrinogen A, protein S alpha and C-reactive protein at early time points in pregnant bitches. In conclusion, the combination of both methods allowed the identification of several altered proteins, and the quantification and description of the concentration patterns for a selection of them during the early stage of dog pregnancy.