B. Kinetics of FadA production in Fn. Fn 12230 and US1 were inoculated to OD600 = 0.1. 10 OD600 units of bacteria were harvested by centrifugation at 12-hr intervals. Bacteria were lysed with 2 mg/ml lysozyme, followed by incubation in 1% sarkosyl at 4° C for 20 minutes, which became the 'total' sample. Lysates were then centrifuged at 100,000 g for 20 minutes and the supernatants and pellets were collected. An aliquot of 5 μl is loaded onto each lane, followed by immunoblot using anti-FadA monoclonal antibody (mAb) 7H7 at 1:4,000 dilution to detect FadA protein in total, supernatant, and pellet fractions. Ponceau staining of lysozyme is shown as a loading control. Pre-FadA and mFadA are pointed by arrows. Note pre-FadA migrates faster than mFadA on SDS-PAGE as previously reported (Han et al., 2005). 