Carbapenemase producing Pseudomonas aeruginosa (CP-PA) is one of are important causative of nosocomial infections and are associated with increase severity of the disease .The increasing carbapenem resistance of P. aeruginosa which have a broad spectrum of antibacterial activity and are used as last resort drugs for the treatment of infections may be mediated by transferable horizontal genes transfer. Rapid detection technique is important to provide early information for appropriate treatment. The aim of this study was to determine specific spectrum to differentiate carbapenemase producing P. aeruginosa (CPPA) from carbapenemase non-producing (CNPPA) and carbapenem susceptible P. aeruginosa (CSPA) by carbapenemase producing activity using a MALDI-TOF MS. Methods: A total of 162 clinical P. aeruginosa strains were composed of 81 carbapenems resistant P. aeruginosa (CRPA) and 81 CSPA isolates. All isolates were routinely identified into species level using biochemical tests and detect CRPA by imipenem and meropenem disk diffusion method. All CRPA isolates were investigated carbapenemase gene by PCR for blaIMP and blaVIM For the CPPA were detected by imipenem hydrolysis assay using MALDI-TOF MS. Results: The CP-PA 39 isolates produced different carbapenemase (IMP and VIM) peaks representing imipenem and degradation products were detected m/z 274, could discriminate between non CPPA were detected m/z 300 by MALDI-TOF MS as follows: 39 of 81 strains were CP-PA, 42 of 81 strains were non-CP-PA. There was a significant relationship between MALDI-TOF MS with PCR (P value < 0.001). The Kappa statistic was 0.877, was excellent agreement between the methods. The sensitivity, specificity, PPV and NPV 90.7%, 97.4%, 97.5% and 90.5% respectively. The MALDI-TOF MS assay was a quick and reliable method can be differentiation to CP-PA from non-CP-PA in routine microbiological diagnosis