Free NET-like DNA structures in CF lung disease. (a) Immunological characterization of free DNA structures in CF airway fluids by CLSM. Upper two panel rows: NET-like DNA structures in induced CF sputum. Blue: DAPI stains DNA-NET backbone. Red: elastase. Lower two panel rows: Blue: DAPI, red: citrullinated histones. Scale bar: 20 μm. (b) Ultrastructure of free NET-like DNA structures. Upper and middle panels: SEM images of CF airway fluids. Arrow marks bacteria entrapped in DNA-NET-like structures. Scale bar: 2 μm; Middle right panel: TEM staining of citrullinated histones in CF airway fluids (sputa). Lower panel: Ultrathin sections of CF airway DNA-NET-like structures. The NETs and the bacterial extracellular polysaccharides are visualized by the ruthenium-red-osmium-tetroxide technique. Bacteria embedded in a dense wickerwork of NETs. Arrowheads mark NETs; asterisk: bacterial extracellular polysaccharide. (c) Upper panel: free DNA structures in CF lung tissue. Red: MPO (as characteristic NET component). Blue: DAPI (DNA). Inlays mark characteristic NET-areas. Lower panel: CF airway NETs in CF BAL fluids. Red: elastase (as characteristic NET component). Blue: DAPI (DNA). (d) Costainings of DAPI, citrullinated histones, and F-actin. Scale bar: 20 μm. (e) Dead/live staining of CF airway fluids (induced sputum). Free DNA and dead bacteria appear red; vital bacteria appear green.