Central venous catheter (CVC)-related thrombosis and infections are frequently occurring complications in patients with hematological malignancies. At present, heparin is most often used as a locking solution. Trisodium citrate (TSC) had been shown to be a very effective antimicrobial catheter locking in hemodialysis patients. We performed a prospective randomized phase III multicenter trial to determine the efficacy of TSC as a locking solution compared to heparin in preventing CVC-related thrombosis and infections in patients with hematological malignancies. Thirty-four episodes of CVC-related bloodstream infections (CVC-BSI) occurred in the 108 patients who were randomized to locking with heparin compared with 35 episodes in the 99 patients who were randomized to locking with TSC (P = 0.654). We did find seven times more CVC-BSI with gram-negative rods in CVCs locked with heparin (P = 0.041). The cumulative incidence of symptomatic thrombosis was 10 % in the heparin group and 5 % in the TSC group (hazard ratio 0.525; 95 % confidence interval 0.182-1.512). This study shows that locking with TSC in patients with hematological malignancies significantly reduced the incidence of CVC-BSI with gram-negative rods. However, the incidence of CVC-BSI with coagulase-negative staphylococcus or CVC-related thrombosis was not reduced by TSC locking.