Background: Detection of anti-SARS-CoV-2 antibodies among people at risk of infection is crucial for understanding both the past transmission of COVID-19 and vulnerability of the population to continuing transmission and, when done serially, the intensity of ongoing transmission over an interval in a community. We aimed to estimate the seroprevalence of COVID-19 in a representative population-based cohort in Iquitos, one of the regions with the highest mortality rates from COVID-19 in Peru, where a devastating number of cases occurred in March, 2020. Methods: We did a population-based study of SARS-CoV-2 transmission in Iquitos at two timepoints: July 13–18, 2020 (baseline), and Aug 13–18, 2020 (1-month follow-up). We obtained a geographically stratified representative sample of the city population using the 2017 census data, which was updated on Jan 20, 2020. We included people who were inhabitants of Iquitos since COVID-19 was identified in Peru (March 6, 2020) or earlier. We excluded people living in institutions, people receiving any pharmacological treatment for COVID-19, people with any contraindication for phlebotomy, and health workers or individuals living with an active health worker. We tested each participant for IgG and IgM anti-SARS-CoV-2 antibodies using the COVID-19 IgG/IgM Rapid Test (Zhejiang Orient Gene Biotech, China). We used survey analysis methods to estimate seroprevalence accounting for the sampling design effect and test performance characteristics. Findings: We identified 726 eligible individuals and enrolled a total of 716 participants (99%), distributed across 40 strata (four districts, two sexes, and five age groups). We excluded ten individuals who: did not have consent from a parent or legal representative (n=3), had moved to Iquitos after March 6, 2020 (n=3), were in transit (n=2), or had respiratory symptoms (n=1). After adjusting for the study sampling effects and sensitivity and specificity of the test, we estimated a seroprevalence of 70% (95% CI 67–73) at baseline and 66% (95% CI 62–70) at 1 month of follow-up, with a test-retest positivity of 65% (95% CI 61–68), and an incidence of new exposures of 2% (95% CI 1–3). We observed significant differences in the seroprevalence between age groups, with participants aged 18–29 years having lower seroprevalence than those aged younger than 12 years (prevalence ratio 0·85 [95% CI 0·73–0·98]; p=0·029). Interpretation: After the first epidemic peak, Iquitos had one of the highest rates of seroprevalence of anti-SARS-CoV-2 antibodies worldwide. Nevertheless, the city experienced a second wave starting in January, 2021, probably due to the emergence of the SARS-CoV-2 P1 variant, which has shown higher transmissibility and reinfection rates. Funding: Dirección Regional de Salud de Loreto (DIRESA), Loreto, Peru. Translation: For the Spanish translation of the abstract see Supplementary Materials section.