Quantitative measurement of the β-lactamase activity of M. catarrhalis and recombinant H. influenzae strains. β-lactamase activity was measured using the chromogenic compound nitrocefin. Bacterial suspensions were mixed with a 250 μg/mL nitrocefin solution and the A486 was immediately measured and recorded as time “0” (open bars). The A486 of the samples was measured again after a 30-min incubation at room temperature (black bars). Panel A: The β-lactamase activity of M. catarrhalis O35E is compared to that of the bro-2 mutant, O35E.Bro, carrying plasmids pWW115, pTS.Bro, and pTS.BroKK. Panel B: The β-lactamase activity of H. influenzae DB117 carrying plasmids pWW115, pTS.Bro, and pTS.BroKK is compared. Sterile broth was used as a negative control in these experiments. The results are expressed as the mean ± standard error A486. Asterisks indicate that the reduction in the β-lactamase activity of strains lacking expression of BRO-2, or expressing a mutated BRO-2 that contains two lysine residues in its signal sequence instead of 2 arginines, is statistically significant (P < 0.05) when compared to bacteria expressing a WT copy of the bro-2 gene.