ProteinReader (PT) is an expert software developed for the analysis of spectra produced with patented in-solution CAF-/CAF+ peptide derivatization reagent. PT is developed as client-server application to analyse MS/MS spectra on MALDI TOF/TOF mass spec. Four main advantages of CAF-/CAF+ technology are: • Allows to capture same peptide in positive and negative mode of operation – additional accuracy control • In positive mode of operation obtaining mostly Y-ions while in negative mode obtaining mostly B-ions - eased de novo readout • No degradation of the sample due to use of CAF-/CAF+ derivatisation reagent • Except unambiguous biotypization (species identification), antibiotic resistance can be easily find by aforementioned method ProteinReader uses two methods for peptide search: database match (NCBInr) and de novo reading. Each peptide can be read with the two spectra, in the positive and the negative ion mode. Testing of more than 200 microorganisms showed positive identification on the species level (strain was also successfully identified if present in NCBInr database). ProteinReader was developed using open source technologies (Java, Eclipse RCP, Linux, Apache Tomcat, Infobright data warehouse) and is designed to work effectively (parallel processing) in a multitier client-server architecture. • CAF-/CAF+ patent: http://www.google.com/patents/EP2529233A2?cl=en • PBF Section for Bioinformatics: http://proteinreader.com/ • IRB Laboratory for system biomedicine: http://www.irb.hr/eng/Research/Divisions-and-Centers/Division-of-Molecular-Medicine/Laboratory-for-system-biomedicine • This work was funded by HRZZ (Croatian Science Foundation) research project “Clinical proteomics of microorganisms”.