Objectives: This study was aimed to assess validation and reliability of knowledge of, attitude toward and practice (KAP) of a Case-mix and Diagnosis Related Group (DRG) system questionnaire. Methods: A sample of 238 health care providers selected conveniently from three public hospitals in Turkey was enrolled in a cross-sectional study from September 1 until November 30, 2012. The mean age was 38.63 years (standard deviation [SD] 10.52), ranging from age 21 to 60 years. More than one-half were males (52.1%), nearly two-fifths were medical doctors (39.9%), one-third were nurses (33.2%), one-sixth were auxiliary staff (16.4%) and the remaining were coders (10.5%). Only one-third (33.6%) of respondents attended a workshop or training program in the Case-mix or DRG system. After examining content validity, factor analysis was conducted, internal consistency of the questionnaire was assessed by Cronbach's alpha estimate, and test-retest reliability was evaluated. Results: The sample adequacy for extraction of the factors was confirmed by the Kaiser-Meyer-Olkin test (0.915) and the Bartlett test (1052). Factor analysis showed three factors, including attitude (36.43%), practice (23.39%) and knowledge (17%), with a total variance of 76.82%. The reliability of each section of the questionnaire was as follows: knowledge (0.963), attitude (0.964) and practice (0.973). Cronbach's alpha total was 0.941, which showed excellent internal consistency. Conclusions: This study demonstrated that the designed questionnaire provided high construct validity and reliability, and could be adequately used to measure KAP among health care staff of the Case-mix and DRG system in Turkey.