Simple SummaryWhen implementing dairy crossbreeding in purebred Holstein (HO) herds, farmers expect to improve the overall herd performance. However, they lack knowledge about how to manage and benefit from the diversity of genetic classes generated by three-breed rotational crossbreeding, which firstly refers to the cohabitation of purebred HO and first- and second-generation crosses (F1 and G2, respectively) within the herd. This study aimed to compare milk production, reproduction and udder health performance of HO, F1 and G2 cows, and to estimate how their combination in different proportions in the herd affects its profitability. We found that HO, F1 and G2 had different and complementary performance profiles, with two main trends. First, HO had higher milk yield, while F1 and G2 crosses had better fertility performance. Second, F1 had win-win trade-offs between milk production, fertility and udder health compared to HO and G2. We showed that HO-F1 or HO-F1-G2 (below 30%) mixed herds could be more profitable than purebred HO or fully crossbred herds with a conventional milk price. These findings can be used for advising purebred HO farmers who wonder about the benefits and the ways of managing the diversity of animal entities generated by the use of dairy crossbreeding in their herds.Using three-breed rotational crossbreeding in a purebred Holstein (HO) herd raises two questions: Do the different genetic classes of cows generated by crossbreeding perform differently? Are there any economic benefits of combining them within a herd? This study aimed at comparing the performance between the different genetic classes resulting from the use of three-breed rotational crossbreeding, and simulating the effect of combining them on herd profitability. Based on a dataset of 14 French commercial dairy herds using three-bred rotational crossbreeding from a HO herd over a 10-year period, we defined three genetic classes according to the theoretical value of heterosis and the percentage of HO genes. We performed linear models and estimated least square means to compare HO cows and the first and second generation of crosses (F1 and G2, respectively) on eight performance characteristics related to milk yield and solids, udder health and fertility. We used these to simulate profitability of five herd compositions differing according to HO, F1 and G2 proportions. We showed that HO, F1 and G2 cows had different and complementary performance profiles. HO had a win-lost trade-off between milk yield and fertility, G2 had the opposite trade-off and F1 had a win-win trade-off. Differences regarding milk solids and udder health were less clear-cut. We highlighted that combining HO with F1 or with both F1 and G2 (below 30%) could be more profitable than using purebred HO or crossbred herds in a conventional milk price scenario. These findings provide evidence on the benefits of functional diversity generated from the use of dairy crossbreeding in dairy herds.