Figure 2.

Quantitative PCR with molecular beacons of nasal brushing, endobronchial brushing, and bronchoalveolar lavage (BAL). Universal primers for bacterial 16S rDNA. The location of brushing is shown on the x axis. The brushes were placed in lysozyme solution and incubated at 65°C for 30 minutes followed by the addition of sodium dodecyl sulfate and proteinase K. The BAL was kept on ice and spun at low speed, followed by separation of fluid from the cell pellet. Aliquots of BAL fluid (200 μl) were then incubated in sodium dodecyl sulfate and proteinase K for 12 hours at 65°C. DNA was extracted from 100,000 BAL cells. In all samples, 2.5% of a 200 μl extract were used in the quantitative PCR reactions.