Background and aims: Knowledge, attitude, and practice (KAP) toward coronavirus disease 2019 (COVID-19) are important factors for adherence to protection strategies. The aim of this study was to construct and validate a questionnaire about the KAP of Iranians regarding COVID-19. Methods: Initially, a questionnaire about the KAP with regard to COVID-19 was constructed in this cross-sectional study. The initial questionnaire was prepared based on a review of the extensive research literature and global and national guidelines. This researcher-made questionnaire was developed using the COVID-19-KAP questionnaire, which was originally used in China in 2020, and the influenza epidemic KAP Questionnaire (2015), the MERS KAP Questionnaire (2015), the H1N1 flu KAP questionnaire (2017), and the WHO KAP questionnaire about COVID-19. The applied questionnaire consists of three parts including knowledge- (n=26), attitude- (n=8), and practice-related (n=11) questions, respectively. Then, the face, content, and construct validity and the reliability of the questionnaire was determined, and analyzes were performed using SPSS v. 26 and AMOS v. 24 software. Results: In the face validity stage, 2 questions with an impact score of less than 1.5 were excluded from the questionnaire. In the content validity check, all questions had a content validity index (I-CVI) of more than 0.7. However, the content validity ratio (CVR) for 6 questions was less than 0.51, and these questions were removed accordingly. The compliance of the CVI with the chance agreement was close to one for all questions, indicating the lack of a chance agreement. In the exploratory factor analysis stage, the Kaiser-Meyer-Olkin (KMO) value of 0.90 was obtained, implying the adequacy of the sample size for factor analysis. The significance of the Bartlett test (chi-square: 5820.06, df: 630, P<0.001) indicated a strong correlation between the questions and the appropriateness of factor analysis. Finally, 34 items in 4 domains remained in the questionnaire. Conclusion: The validity and reliability of this questionnaire were confirmed based on the findings. This Farsi KAP questionnaire can be used to assess the level of knowledge, attitude, and practice of people about COVID-19.