In vivo T-cell depletion using anti-thymocyte globulin (ATG) is widely used in allogeneic hematopoietic stem cell transplantation (HSCT) for prophylaxis of GvHD. We investigated the influence of thymoglobulin dose (an ATG) on GvHD following matched sibling donor (MSD) HSCT with a busulfan and fludarabine preparative regimen. Medical records of 180 patients who received MSD HSCT with a conditioning regimen of busulfan, fludarabine, and ATG (BuFluATG) were reviewed retrospectively. The median age was 53 years (range 18-68). Initial diagnoses were acute myeloid leukemia (73.3%) and myelodysplastic syndrome (26.7%). Forty-four and 68 patients (24.4 and 37.7%) experienced acute and chronic GvHD of any grade, respectively. High-dose (⩾4.5 mg/kg) ATG was independently associated with decreased risk of acute GvHD (hazard ratio=0.36, 95% confidence interval (CI): 0.15-0.84, P=0.019) compared to low-dose ATG (<4.5 mg/kg). Although ATG dose was associated with the risk of acute GvHD, it was not associated with the risk of chronic GvHD in our study. A higher dose (⩾4.5 mg/kg) of ATG decreases the risk of acute GvHD but had no significant impact on disease-free survival in MSD HSCT patients conditioned with BuFluATG. The optimal dose of ATG should be further investigated in a large prospective study context.