The invention provides a method for detecting the content of selenium in cigarette mainstream smoke. The method comprises the steps: collecting total particular matter selenium in the cigarette mainstream smoke in an electrostatic collecting manner, simultaneously collecting gas-phase selenium in the cigarette mainstream smoke by adopting an acidified ethanol solution, and measuring the content of selenium in the cigarette mainstream smoke by adopting an inductively coupled plasma mass spectrometry. The invention further provides a method for detecting the content of the gas-phase selenium in the cigarette mainstream smoke and application of the gas-phase selenium in the cigarette mainstream smoke. The method for detecting the content of selenium in the cigarette mainstream smoke has good reproducibility and recovery rate, and is beneficial for measuring the content of selenium in the cigarette mainstream smoke effectively and quantitatively by relative technical staff in the tobacco industry; and a strong technical support is provided for evaluating perniciousness of selenium in the cigarette mainstream smoke in the tobacco industry comprehensively and objectively.