Background: Non-communicable diseases (NCDs) are the leading cause of death worldwide, with a disproportionally rising burden among low- and middle-income populations. While preventable risk factors highly contribute to this burden, population-based studies assessing these factors and the health status of these populations, are scarce. Methods: The Khuzestan Comprehensive Health Study (KCHS)—a cross sectional study—was conducted between 2016-2019, including 30,506 Iranians aged 20 to 65 years, from 27 counties of Khuzestan province, southwest of Iran. KCHS aimed to provide a comprehensive health overview by investigating the prevalence and risk factors of NCDs and psychological disorders, along with viral hepatitis as a common communicable disease. Upon registration, 15 mL of blood and anthropometric measurements were obtained from participants. Afterwards, several interviewer-administered questionnaires were completed to gather data on demographics, socioeconomic status, sleep quality, physical activity, lifestyle habits, nutrition, and medical history. Results: The mean ± SD age of participants was 41.7 ± 11.9 years. The majority were female (64.3%), of the Arab ethnicity (49%), married (83%), and urban residents (73.1%). About 70% had an educational level below high school diploma. Overall, 10.8%, 5.2%, and 2.8% of participants had used cigarettes, hookah, and drugs at least once in their lifetime, respectively. While body mass index and serum cholesterol levels were higher in females, blood pressure was higher in males (P<0.001). Conclusion: KCHS assessed many aspects of health in the Khuzestan province. In addition to develop a biobank along with a comprehensive dataset, KCHS will serve as a valuable infrastructure for future research.