Background: The Directly Observed Treatment Short course Strategy was launched in 1994 to address the problem of Tuberculosis globally. It consists of five components: - Sustained political commitment, access to quality-assured sputum microscopy, Standardized short- course chemotherapy for all cases of Tuberculosis under proper case management conditions, including direct observation of treatment. Uninterrupted supply of quality-assured drugs, and recording and reporting system enabling outcome assessment of all patients and assessment of overall programme performance. Objective: To assess quality of directly observed treatment short-course of tuberculosis service and its associated factors in Gondar Town Administration, northwest Ethiopia. Methods: Institutional based cross-sectional mixed quantitative and qualitative study design was used. All health facilities and all records meeting eligibility criteria were used to collect data. Data were collected by using document review and checklist from records and in-depth interview from health professionals. The data was coded & entered in to Epi Info version 7. Then the data were exported to SPSS version 20 for analysis. Bivariable & Multivariable logistic regression was employed to see the existence of association between quality of directly observed treatment short-course of tuberculosis & independent variable. Results: The results of this study showed that Input, process and output quality parameters were 73.4%, 70.95%, and 67.5% respectively, with the overall quality of 70.4%. Variables such as Patients didn’t participated in part of decision making [AOR=2.32 (95% CI 1.31- 4.11)], patient privacy [AOR=1.85 (95% CI 1.01–3.37)], Completeness of information on Tuberculosis registry and [AOR =2.61(95% CI 1.13-5.98)], time taken to reach to health facility [AOR=2.13 (95% CI 1.29-4.21)] were significantly associate with quality of Directly Observed Treatment Short course. Conclusion and recommendation: Input, process and output qualities of a program were poor in relative to the 100% World Health Organization requirement. In our study the total quality of Directly Observed Treatment Short course Strategy was low. There is structural problem & irregular supervision. Patients were dissatisfied regarding to cleanliness, comfortableness of waiting area, duration of waiting time, and adequacy of working hours of Tuberculosis clinic which can lead to patients’ failure to adhere to treatment, service rejection and program failure. These phenomena indicated that the problem need due attention at each level by the concerned bodies to alleviate the problems and improve the quality. Key words: DOTS, Gondar town Administration zone. Quality