Background: Spiritual well-being is an important issue in health sciences, hence the need for validated instruments to assess this aspect of health in the Iranian population. The aim of the current study was to determine the validity of the Persian versions of 2 most common measures of spiritual health (Spiritual Well-Being Questionnaire [SWBQ] or Spiritual Health and Life-Orientation Measure [SHALOM] and Spiritual Well-Being Scale [SWBS]). Methods: This was a cross-sectional study via a convenience sampling method in Iran University of Medical Sciences with 170 participants aged above 18 years comprising students, teachers, and administrative staff and managers. The study was conducted from September 7, 2014 to September 20, 2015 in Tehran. Four questionnaires, namely the SWBQ, SWBS, General Health Questionnaire (GHQ-12), and Oxford Happiness Questionnaire (OHQ), were used. Statistical analysis was done using SPSS 18 and LISREL (version 8.2). Cronbach’s alpha, intra-class correlation coefficient, Pearson correlation, and confirmatory factor analysis were employed to assess the validity and reliability of the questionnaires.Results: Cronbach’s alpha for the SWBQ and the SWBS was greater than 0.85. The repeatability of both questionnaires was between 0.88 and 0.98. The Pearson correlation for the SWBQ and the SWBS ranged from 0.33 to 0.53; and all the correlations were significant. The respondents who indicated a higher spiritual well-being also reported better general health and happiness.Conclusion: The Persian versions of the SWBS and the SWBQ have good reliability, repeatability, and validity to assess spiritual health in the Iranian population.