Ha-toxin QuantificationAlpha-toxin production by the CA-MRSA and HA-MRSA strains was assessed by indicates of a sandwich ELISA. In brief, the wells of microtiter plates have been coated with an anti-alpha-toxin murine monoclonal antibody (kindly offered by GSK Biologicals) in PBS overnight at space temperature. The unbound monoclonal antibody was washed out twice having a blocking answer of PBS-Tween (0.05 ) and milk (five g/L), followed by incubation with the blocking answer for 1 h at area temperature. Standard dilutions of recombinant alpha-toxin and 24 h bacterial culture supernatant had been added to duplicate wells, incubated for 1 h at 37uC, and washed three times, followed by the addition of aPLOS One | www.plosone.orgSupporting InformationTable S1 A comparison of distinct lineages of CA-MRSA and HA-MRSA with respect to cytotoxicity towardCA-MRSA PSMs Kill Osteoblastshuman osteoblasts, intracellular survival and alphatoxin production.HEPES Technical Information (PDF)C. Courtier, C. Gardon, and C. Bouveyron for professional technical enable; P.J. Bondon for performing the LDH assays; and T. Henry for beneficial suggestions.Author ContributionsConceived and made the experiments: JPR FV FL. Performed the experiments: JPR STA AS YL JR FC CB YB MB. Analyzed the data: JPR. Contributed reagents/materials/analysis tools: TF Bad ST GL JE FV. Wrote the paper: JPR FV JE FL.AcknowledgmentsThe authors would like to thank the bacteriologists throughout the globe who sent CA-MRSA and HA-MRSA strains for the FNRCS; C. Hayraud,
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