Introduction: The use of the matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry has shown to be effective and fast in some clinical specimens for the identification of colonizing microorganisms. The objective of the study was to analyze the validity values for the prediction of colonization and catheter-related bloodstream infection (C-RBSI) of the MALDI-TOF mass spectrometry performed at all intravascular catheters that arrived in the microbiology laboratory., Methods: Catheter tips (after performing the roll-plate technique) were tested by MALDI-TOF mass spectrometry during a period of 3-months. The gold standard for colonization and C-RBSI were, respectively: the presence of ≥15cfu/plate in the catheter tip culture; and the isolation of the same microorganism(s) in blood cultures as well as in the colonized catheter (during the 7days before or after catheter withdrawal)., Results: A total of 182 intravascular catheters were collected. The overall colonization rate detected by roll-plate technique and MAL-TOF mass spectrometry was 31.9% and 32.4%, respectively. Overall, there were 33 (18.1%) episodes of C-RBSI. The validity values of the MALDI-TOF mass spectrometry for the identification of colonization and C-RBSI were, respectively: sensitivity (69.0%/66.7%), specificity (84.7%/75.2%), positive predictive value (65.6%/36.1%), and negative predictive value (86.8%/92.6%). Conclusion MALDI-TOF mass spectrometry could be an alternative diagnostic tool for ruling out C-RBSI. However, despite it showing to be faster than conventional culture, future studies are required in order to improve the pre-analytical process., (Copyright © 2013 Elsevier España, S.L. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.)