Identifying the drivers of wildlife immunity is critical for assessing stressor impacts and zoonotic risks. However, such studies are limited by the logistical challenges of wildlife research and the lack of species-specific reagents. We here optimize flow cytometry, typically confined to laboratory settings, to field settings to profile cellular immunity with small blood volumes and extended sample holding times. We apply these methods to analyze immune cell seasonality in migratory Mexican free-tailed bats (Tadarida brasiliensis). We confirmed four antibodies recognizing CD3, CD79a, MHC II, and CD11b that were originally validated in Egyptian fruit bats (Rousettus aegyptiacus), allowing us to quantify T and B cells, macrophages, and neutrophils, respectively. Flow cytometry outperformed traditional hematology in quantifying leukocyte profiles and revealed pronounced seasonality in cellular immunity. Innate cells dominated after spring migratory arrival, whereas adaptive cells dominated during pregnancy and before fall migration. Neutrophils and B cells peaked during pregnancy and after pup weaning, respectively, while macrophages and T cells had no seasonality. Females had more B cells than males, but did not differ in other cell proportions. Our findings lay the groundwork for applying flow cytometry to field studies of wildlife and provide important insights into the seasonality of wild bat immunity.