These normalized results were used to calculate the fold change expression of ureC during growth in CDM plus sputum
compared to CDM alone. BioRad iQ5 software was used to analyze data. Enzyme-linked immunosorbent assay (ELISA) Eighteen pre and post exacerbation serum pairs from adults with COPD followed in the find more COPD Study Clinic were subjected to ELISA to detect the development of new IgG antibodies in serum to urease C . The change in antibody level from pre-exacerbation to post-exacerbation samples was calculated using the following formula: % change = [( post OD - pre OD )/pre OD] × 100. Paired pre-exacerbation and post-exacerbation samples were always tested in the same assay. The cutoff value for a significant percentage change between pre-exacerbation and post-exacerbation serum IgG levels was determined by studying 8 control pairs of serum samples obtained 2 months apart (the same time interval for the experimental samples) from patients who were clinically stable and who had negative sputum cultures for H. influenzae as described previously [42, 43, 48, 63]. Susceptibility of H. influenzae to acid H. influenzae wild type and mutant strains were grown in broth to log phase, harvested by centrifugation and suspended to a concentration of ~107 colony forming units/ml in PBS
adjusted to varying pH. learn more Cells were incubated in the Apoptosis inhibitor presence or absence of urea (50 mM or 100 mM) and dilutions of bacteria were plated at time 0 and at 30 min. Bacteria were counted after overnight incubation on chocolate agar. Acknowledgements and Funding This work was supported by National Institutes of Health grant
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