Simple SummaryIn autumn calving dairy herds cattle must conceive within a restricted timeframe to maintain the seasonality of calving. Cattle not observed in estrus prior to the mating start date are frequently treated with a single prostaglandin injection even though its relative effectiveness is unknown. The perceived cost of other treatments, such as a modified Ovsynch protocol, may be a reason why they are less favored. This study compares the reproductive and economic outcomes associated with treating cows not observed in estrus prior to breeding with a modified Ovsynch protocol, in comparison to a single prostaglandin injection. The main inputs influencing profitability are identified, and differences in dry-off strategy are considered. From the analyses, cows treated with a modified Ovsynch protocol had a greater likelihood of conception at the beginning of, and throughout, the breeding season. Differences in barren rate and the amount of milk produced were the main variables affecting profitability, the latter strongly influenced by drying-off policy. Farmers and advisors should consider the relative performance of their breeding strategies as well as their own management policies to optimize reproductive and economic performance.In autumn calving dairy herds, treatment of cattle not observed in estrus prior to the breeding season is common. Routinely, a single prostaglandin or a modified Ovsynch (MOFT) protocol are used—without evidence of their relative effectiveness. This study compares the effects on conception, associated timing, and profitability of administering cows with prostaglandin or MOFT treatment. A hundred and ninety-two Holstein-Friesian cows from three herds without an observed estrus within 28-days before mating start date were randomly treated with d-cloprostenol (PGOD) or an 8-day MOFT protocol. The association of treatment and calving-breeding start-date interval (CBSI) on the risk of conception were investigated. Partial budget, sensitivity analysis, and Monte Carlo simulation was used to assess economic performance, identify critical input variables, and explore the effects of input uncertainties on model output. There was a significant association between MOFT treatment and conception during 21 and 84 days after mating start date, compared to PGOD. MOFT treatment was associated with a mean net benefit of £58.21 (sd £19.42) and £27.29 (sd £17.75) per cow for herds with a fixed or variable dry-off date, respectively. The relative profitability of an MOFT protocol is dependent on its effects on barren rate and herd dry-off strategy.