We evaluated the incidence of GVHD, risk factors and the impact of graft composition on acute GVHD (aGVHD) in 92 children who underwent BMT for thalassemia following busulfan/cyclophosphamide (BUCY)-based conditioning regimens and GVHD prophylaxis with CSA/short-MTX and methylprednisolone. The incidence of grade 2-4 and 3-4 aGVHD was 35% (95% confidence interval (CI) 25-44) and 9% (95% CI 4-16), respectively. We found that CD3 þ and CD34 þ cell doses above the median were associated with high incidence of grade 2-4 aGVHD (49 vs 20%, P ¼ 0.005 and 46 vs 23%, P ¼ 0.021, respectively). In multivariate analysis, high CD3 þ (hazard ratio (HR) 4.6; 95% CI 1.4-14.7; P ¼ 0.010) and CD34 þ (HR 4.3; 95% CI 1.4-12.7; P ¼ 0.011) cell doses were associated with grade 2-4 aGVHD. We further examined the effect of CD3 þ and CD34 þ cell doses on aGVHD using quartile cutoff points and found a minimum threshold for CD3 þ (38 Â 10 6 /kg) and CD34 þ (4 Â 10 6 /kg) cells above which the incidence of grade 2-4 aGVHD is significantly increased. This study shows for the first time a positive correlation between the number of CD3 þ and CD34 þ cells and aGVHD in children receiving sibling BMT, and indicates that using tailored and more intensive post transplant immunosuppression may permit to better control aGVHD.