Introduction: Transmission of Trypanosoma cruzi through blood transfusions represents the second most frequent mechanism after the vectorial transmission. Therefore, it is necessary to identify some socio epidemiological characteristics that allow predicting the Trypanosoma cruzi infection on the donors. Objective: To determine the prevalence of anti-Trypanosoma cruzi and risk factors for seropositivity in voluntary blood donors at the Hemocentro y Unidad de Aferesis de Valledupar, Colombia. Materials and methods: A descriptive cross-sectional study was conducted in 170 volunteer blood donors recruited between 2013 and 2014. Detection of IgG antibodies anti-Trypanosoma cruzi was performed by serological tests. The inquiry about the predictors of seropositivity for Trypanosoma cruzi was made through a socioepidemiological questionnaire, followed by a bivariate analysis and logistic regression. Results: The overall prevalence of infection for Trypanosoma cruzi was of 0.14%. The variables that predict the seropositivity for Trypanosoma cruzi of blood donors were the presence of the vector in the house, being bitten by the vector at least once in life and dwell in a house with rammed earth or bajareque during childhood. Conclusions: Although the prevalence of anti-Trypanosoma cruzi antibodies was low in blood donors of Valledupar, Colombia, this study reveals the importance of incorporating questions in the questionnaire of donor selection to warn the professionals of blood bank about a possible infection with the parasite, and therefore, optimize the screening of blood units and their classification as unsuitable for transfusion.