Objective To design a questionnaire for sub-health status of college students and analyze its reliability and validity. Methods Based on literature review and expert consultation, an initial questionnaire with 52 items was formed. A random sample including 1200 students was conducted a questionnaire survey. Formal questionnaire was formed through expert consultation, project analysis and exploratory factor analysis. Factor analysis and reliability analysis were used to analyze the validity, and the reliability. Results A total of 1186 valid questionnaires were callback(98.83%). The formal questionnaire contained 42 items and 3 dimensions with a cumulative variance contribution rate of 59.06%. The Cronbach′s α coefficient equaled to 0.834, the split half reliability equaled to 0.735. The body dimension contained 23 items and 7 common factors, with a cumulative variance contribution rate of 62.80%, and the Cronbach′s α coefficient fluctuated between 0.660 and 0.819, the split half reliability fluctuated between 0.549 and 0.734. The social adaptation dimension contained 10 items and 2 common factors with a cumulative variance contribution rate of 51.20%, the Cronbach′s α coefficients were 0.799 and 0.673, the split half reliability were 0.735 and 0.648. The psychological dimension contained 9 items and 1 common factor with a cumulative variance contribution rate of 49.32%, the Cronbach′s α coefficient was 0.870, the split half reliability was 0.817. Conclusion The questionnaire of sub-health status for college students had good reliability and validity. It could be used as an effective tool to evaluate the sub-health status of college students.