Staphylococcal Isolates Identification via MALDI-TOF-MS

All isolates were initially screened using conventional methods including microscopy (Gram film), catalase and coagulase testing. Prolex™ staph latex kits (ProLab Diagnostics, Neston, UK) were used to identify CNS and S. aureus. All potential staphylococcal isolates were then fully identified using a Matrix-assisted laser desorption ionization time flight mass-spectroscopy (MALDI-TOF-MS, Microflex LT, Bruker Daltonics, Coventry, UK) in a positive linear mode (2000–20,000 m/z range). Samples were prepared as described previously (Mkrtchyan et al., 2013 (link)). In brief, 3–5 colonies of fresh cultures were added into 300 μl distilled water and mixed with 900 μl absolute ethanol. The suspension was centrifuged for 2 min at 13,000 × g and the pellets were re-suspended in 25 μl of 70% formic acid and then mixed with 25 μl pure acetonitrile. The mixture was centrifuge for 2 min at 13,000 × g. One microliter aliquots of the obtained supernatant were spotted on the MALDI target plate and overlaid with 1 μl of α-Cyano-4-hydroxycinnamic acid (HCCA) matrix (Bruker Daltonics, Coventry, UK). The resulting spectra for each isolate was analyzed by MALDI-Biotyper 3.0 software (Bruker Daltonics, Coventry, UK). Escherichia coli DH5α (Bruker Daltonics, Coventry, UK) was used as a standard for calibration and quality control.