Timely diagnosis of pregnancy and pregnancy losses is economically important. A commercially available pregnancy-associated glycoprotein (PAG) milk assay (Idexx Laboratories Inc.) is offered through routine Dairy Herd Improvement (DHI) testing for diagnosis of pregnancy. The objective of this observational study was to describe the relationship between PAG at various stages of gestation and the likelihood of successful calving. The hypothesis was that higher PAG levels would be associated with successful calving. Data were collected from CanWest DHI for all cows with a milk PAG assay between January 1 and May 31, 2013. The PAG milk assay result is reported as a relative PAG level (S-N value). Cows that tested pregnant (PAG >0.25 according to the manufacturer’s cut-point; 6196 cows in 967 herds) were included in this analysis. A dichotomous outcome of calving between 270 and 290 d after the relevant insemination was determined for each cow with 5139 successful outcomes. There was a significant interaction between days in gestation (DIG) at time of PAG assay and PAG, so the data were stratified by DIG. Milk PAG levels increased after breeding, decreased at 45–75 DIG, then increased through the remainder of gestation. For cows ≤45 DIG (n = 793) increasing PAG level was associated with an increased likelihood of calving (P = 0.01). For cows > 45 and ≤75 DIG (n = 1653) relative PAG levels (P = 0.001) and linear score (P = 0.01) were negatively associated with the outcome, while test-day milk yield was positively associated with the outcome (P = 0.01). For cows >75 DIG (n = 3750) relative PAG level (P = 0.001), and milk yield (P = 0.005) were positively associated with full-term pregnancy; linear score was negatively associated with the outcome (P = 0.05). These results indicate that while higher PAG levels are positively associated with a positive calving outcome in general, a decrease in PAG level around 45–75 d in gestation was associated with a successful pregnancy outcome.