Background: Existing study quality and risk of bias lists for observational studies have important disadvantages. For this reason, a comprehensive widely applicable quality assessment tool for observational studies was developed. Methods: Criteria from three quality lists were merged into a new quality assessment tool: the observational study quality evaluation (OSQE). OSQE consists of a cohort, case-control, and cross-sectional version. Results: The OSQE cohort, the OSQE case-control, and the OSQE cross-sectional version include all items applicable to that type of study, for example, the representativeness of the study population, the validity of the independent and dependent variables, and the statistical methods used. Before scoring the OSQE, the rater is asked to define how to score items, in detail. A study can obtain a star for each item. Each item also has a veto cell. This cell can be checked when poor quality with respect to that specific item results in a low quality of the study despite stars on other items. Although stars add to a sum score, the comment field is the most important part of the OSQE. Conclusion: The OSQE presented in the current article provides a short, comprehensive, and widely applicable list to assess study quality and therewith risk of bias.