Biofilm-regulated protein expression. SYPRO® Ruby stained 2-D gels of total protein extracts from flowing biofilm (A) and planktonic (B) cells of S. Typhimurium grown in CFA medium at 25°C for 72 h. Spots circled in red were excised from the gels and identified by mass spectrometry and peptide mass fingerprinting. (C) Magnification of 2-D gels comparing expression of FlgK (I), DppA (II), and YggE (III), which were all more highly expressed in the biofilm cells than in planktonic cells. (D) Western immunoblot of total protein extracts of mature S. Typhimurium biofilm (B) and planktonic (P) cells both grown in CFA medium for 72 h. Protein extracts were separated on a denaturing SDS-PAGE gel and probed with anti-FliC monoclonal antibody. Densitometric analysis showed that FliC was 3.4 fold induced in the biofilm compared with planktonic cells.