The airway surface liquid (ASL) that lines the airway surface epithelium plays a major role in airway antibacterial defense and mucociliary transport efficiency, two key factors in cystic fibrosis (CF) disease. A major difficulty is to collect ASL in native conditions without stimulation or alteration of the underlying airway epithelium. Using a cryoprobe specifically adapted to collect native ASL from the tracheal mouse surface, we analyzed by X-ray microanalysis the complete ASL and plasma ion content in Cftrtm1Hgu/Cftrtm1Hgu mice compared with that in control littermates. ASL ion content from eight Cftrtm1Hgu/Cftrtm1Hgu mice and eight control littermates did not appear significantly different. The mean (±SE) concentrations were 2,352 ± 367 and 2,058 ± 401 mmol/kg dry weight for Na, 1,659 ± 272 and 1,448 ± 281 mmol/kg dry weight for Cl, 357 ± 57 and 337 ± 38 mmol/kg dry weight for S, 1,066 ± 220 and 787 ± 182 mmol/kg dry weight for K, 400 ± 82 and 301 ± 58 mmol/kg dry weight for Ca, 105 ± 31 and 105 ± 20 mmol/kg dry weight for Mg, 33 ± 15 and 29 ± 9 mmol/kg dry weight for P in non-CF and CF mice, respectively. This cryotechnique appears to be a promising technique for analyzing the complete elemental composition of native ASL in CF and non-CF tissues.