Mucus-penetration assay. (a) OC/DNA and C/DNA nanocomplexes. Lane 1: mucus only, lane 2: mucus + naked DNA, lanes 3–7: mucus + OC/DNA nanocomplexes at N/P ratios of 2, 5, 10, 20, and 30, respectively, lanes 8–12: mucus + C/DNA nanocomplexes at N/P ratios of 2, 5, 10, 20, and 30, respectively. (b) mPEG–OC/DNA and mPEG–C/DNA nanocomplexes. Lane 1: mucus only, lane 2: mucus + naked DNA, lanes 3–7: mucus + mPEG–OC/DNA nanocomplexes at N/P ratios of 2, 5, 10, 20, and 30, respectively, lanes 8–12: mucus + mPEG–C/DNA nanocomplexes at N/P ratios of 2, 5, 10, 20, and 30, respectively. (c) Illustration of mucus permeation assay for chitosan/DNA nanocomplexes. (d) Illustration of mucus permeation assay for PEGylated chitosan/DNA nanocomplexes.