Introduction: Chagas disease is a public health problem in Latin America. Even though vector-borne infection is the most important transmission mode for this disease, other modes such as transfusions require evaluation. Objective: To describe the prevalence of T. cruzi infection in multitransfused patients. Materials and methods: We detected IgG antibodies against T. cruzi by two immunoassays in samples from multitransfused patients in four hospitals located in Bogotá and Medellín, Colombia. We analyzed the association with known risk factors, and we calculated the odds ratios (OR) with 95% confidence intervals using Stata 11™ statistical software. Results: In total, 479 samples were tested. Overall, T. cruzi antibody prevalence was 1.88% (nine patients). Five were onco-hematological patients, two were hemodialyzed, one had thalassemia, and one had suffered acute blood loss. We found no hemophilia patients. There was no association between known risk factors for transfusion-transmitted infection (such as the number of transfusion events, number of blood units and type of blood component) and the presence of anti-T. cruzi antibodies in this study. Only the hepatitis C virus infection showed a positive association with the presence of anti-T. cruzi antibodies (OR=5.68, 95% CI: 1.36-23.63). Conclusions: The results of this study showed a low frequency of T. cruzi infection in multitransfused patients, suggesting that the risk of transfusion infection in Colombia is low. Known risk factors for transfusion-related infection were not associated with the presence of anti-T. cruzi antibodies.