Abstract Context The effect of gender-affirming hormone therapy (HT) on erythropoiesis is an area of priority in transgender health research. Objective To compare changes in hematologic parameters and rates of erythrocytosis and anemia among transgender people to those of cisgender controls. Design Longitudinal observational study. Participants and Setting We compared 559 transfeminine (TF) and 424 transmasculine (TM) people enrolled in 3 integrated health care systems to matched cisgender referents. Interventions and Outcome Hormone therapy receipt was ascertained from filled prescriptions. Hemoglobin (Hb) and hematocrit (Hct) levels were examined from the first blood test to HT initiation, and from the start of HT to the most recent blood test. Rates of erythrocytosis and anemia in transgender participants and referents were compared by calculating adjusted hazard ratios and 95% confidence intervals (CI). Results In the TF group, there was a downward trend for both Hb and Hct. The corresponding changes in the TM cohort were in the opposite direction. TM study participants experienced a 7-fold higher rate (95% CI: 4.1–13.4) of erythrocytosis relative to matched cisgender males, and an 83-fold higher rate (95% CI: 36.1–191.2) compared to cisgender females. The corresponding rates for anemia were elevated in TF subjects but primarily relative to cisgender males (hazard ratio 5.9; 95% CI: 4.6–7.5). Conclusions Our results support previous recommendations that hematological parameters of transgender people receiving HT should be interpreted based on their affirmed gender, rather than their sex documented at birth. The clinical significance of erythrocytosis following testosterone therapy, as well as anemia following feminizing HT, requires further investigation.