SDS-PAGE of secreted proteins (A, C) and Western blot for detection of SspA and Hla (B), in culture supernatant of USA300 after growth for 18–24 h in the presence of C16 (A) or C18 (C) fatty acids.

Cultures were grown with the indicated amounts of C16∶1▵6 (sapienic acid), C16∶1▵9 (palmitoleic acid), C16∶0 (palmitic acid), C18∶2 (linoleic acid), C18∶1 (oleic acid), C18∶3 (linolenic acid) or C18∶0 (stearic acid) fatty acids. Proteins in the cell-free culture supernatant were precipitated in ice-cold TCA, and after solubilization in SDS-PAGE reducing buffer, protein equivalent to 2.0 OD₆₀₀ units of culture supernatant was loaded in each lane (A and C). For Western blot (C), 0.02 OD₆₀₀ units of cell free culture supernatant were subjected directly to SDS-PAGE, prior to detection with specific antisera (see Materials and Methods).