Figure 5

Western blotting of salivary MUC5B and MUC7 from HIV positive individuals with different CD4 counts. Lane 1, MUC5B from patients with CD4 > 400, lane 2, MUC5B from patients with CD4 200-400, lane 3, MUC5B from patients with CD4 < 200, lane 4, MUC5B from HIV negative individual, lane 5, crude saliva (positive control), lane 6, MUC7 (negative control), lane 7, MUC7 from patients with CD4 > 400, lane 8, MUC7 from patients with CD4 200-400, lane 9, MUC7 from patients with CD4 < 200, lane 10, MUC7 from HIV negative individuals, lane 11, crude saliva (positive control) and lane 12, MUC5B (negative control) were separated by a 1% agarose gel and transferred to nitrocellulose membrane. Following overnight blocking, the membranes were incubated for 2 h with rabbit anti-MUC5B polyclonal (lanes 1-6) and mouse anti-MUC7 monoclonal (lanes 7-12) antibodies diluted in 5% (m/v) low fat milk powder in TBST at 1 in 2000 (rabbit anti-MUC5B) and 1 in 1000 (mouse anti-MUC7). Membranes were then washed 3 × 5 min with TBST and incubated for 1 h with HRPO linked goat anti-rabbit (lanes 1-6) and goat anti-mouse (lanes 7-12) secondary antibodies diluted in 5% (m/v) low fat milk powder in TBST at dilutions of 1 in 5000 and 1 in 1500 respectively. After another TBST wash (3 × 5 min), bands were detected using an ECL detection kit.