Background: As the COVID-19 pandemic unfolded, rapid case increase was observed in multiple cities in Iran. However, in the absence of seroprevalence surveys, the true infection rate remains unknown. In this population-based study we assessed the seroprevalence of SARS-CoV-2 antibodies in eighteen cities of Iran.Methods: We randomly selected and invited study participants from the general population (N = 3,547) and occupations with high risk of COVID-19 exposure, defined as high-risk population (e.g., supermarket employees) (N = 5,391), in eighteen cities of Iran. SARS-CoV-2 ELISA kits were used to detect antibody against COVID-19. Crude, population weight adjusted, and test performance adjusted seroprevalence rates were estimated.Findings: The population weight adjusted and test performance adjusted prevalence rates of antibody seropositivity in general population were 13·1% (95% CI 11·6-14·8%) and 18·5% (95% CI 16·1-21·3%), respectively. The population-weighted seroprevalence estimate implies that 3,290,633 (95% CI 2,907185-3,709,167) individuals, from the eighteen included cities in this study, were infected by end of April 2020.The overall prevalence rate was higher among individuals aged ≥ 60 years (32·0%, 95% CI 23·9-40·8%) and with comorbidity condition (23·7%, 95% CI 18·5-28·8%). The estimated seroprevalence of SARS-CoV-2 antibodies varied greatly by city and the highest population test-adjusted prevalence rates were in Rasht 78·1% (95% CI 58·3-98·3%) and Qom (66·5%, 95% CI 39·9-95·4%) cities. The test-adjusted prevalence did not differ between low and high-risk populations and was about 20.0%.Interpretations: The findings of this study imply that prevalence of seropositivity is likely much higher than the reported prevalence rates based on confirmed COVID-19 cases in Iran. Despite the high seroprevalence rates in a few cities, the low overall prevalence estimates indicate that a large proportion of population is still susceptible to the virus. The similar seroprevalence estimates between low and high-risk occupations might be an indicator of inadequate or low adherence to infection control measures among general population.Funding Statement: Iranian Ministry of Health and Medical Education COVID-19 Grant (number 99-1-97-47964).Declaration of Interests: None to disclose.Ethics Approval Statement: Ethics approval for this study was granted by Vice-Chancellor in Research Affairs-Tehran University of Medical Sciences (IR. TUMS.VCR.REC.1399.308)