Fig. 1.

Serum opsonization of a Psl-deficient mutant of P. aeruginosa increases the oxidative burst by human neutrophils. A. Immunoblot showing expression of Psl from EPS extracts derived from strains PAO1, WFPA800 and WFPA801 (Byrd et al., 2009). These extracts were blotted on a nitrocellulose membrane, probed with anti-Psl antiserum and detected by chemiluminescence using a HRP-conjugated secondary antibody. B. Kinetic analysis of the chemiluminescence response (measurement of oxidative burst) of neutrophils generated against pre-opsonized Psl variant strains of P. aeruginosa. Each experiment was performed in quadruplet wells (n = 3 with 10% fresh normal human serum opsonization). RLU, relative luminescence unit. C. Oxidative burst response of neutrophils induced by Psl variant strains of P. aeruginosa under fresh serum opsonized (NHS), heat-inactivated serum (HIS)-opsonized and non-opsonized conditions. Bacteria were incubated with serum or no serum, washed and then incubated with neutrophils (moi 50:1) at 37°C for 25 min in presence of luminol and the chemiluminescence response of neutrophils was measured. Each experiment was performed in quadruplet wells (n = 5 for 10% NHS opsonization, HIS opsonization and non-opsonization). Values are mean ± SEM for five experiments. A statistical analysis was carried out using an unpaired two-tailed student’s t-test to compare the chemiluminescence response of neutrophils induced by HIS, opsonized and unopsonized bacteria. The asterisks indicate P-values (*P ≤ 0.05; **P ≤ 0.005). RLU, relative luminescence unit.