Conventional sputum cytology can be used for the detection of lung cancer, but has shown a low yield in prospective screening trials. This review focuses on the technical aspects relevant to the outcome of DNA and image analysis in sputum. Published articles are discussed in the light of the technical background. Recent developments in DNA analysis and nuclear image analysis show a clear potential to improve or refine diagnosis beyond that achieved with conventional sputum cytology examination. The challenge for future studies in DNA and nuclear analysis of sputum is to ensure high levels of quality control and to confirm these initial encouraging results.