Unprofessional behaviors in medical school predict high stakes consequences for practicing physicians, yet little is known about specific behaviors associated with professionalism during residency. To identify behaviors that distinguish highly professional residents from their peers. Comparative study of 148 first-year internal medicine residents at Mayo Clinic from July 1, 2004, through June 30, 2007. Professionalism as determined by multiple observation-based assessments by peers, senior residents, faculty, medical students, and nonphysician professionals over 1 year. Highly professional residents were defined as those who received a total professionalism score at the 80th percentile or higher of observation-based assessments on a 5-point scale (1, needs improvement; 5, exceptional). They were compared with residents who received professionalism scores below the 80th percentile according to In-Training Examination (ITE) scores, Mini-Clinical Evaluation Exercise (mini-CEX) scores, conscientious behaviors (percentage of completed evaluations and conference attendance), and receipt of a warning or probation from the residency program. The median total professionalism score among highly professional residents was 4.39 (interquartile range [IQR], 4.32-4.44) vs 4.07 (IQR, 3.91-4.17) among comparison residents. Highly professional residents achieved higher median scores on the ITE (65.5; IQR, 60.5-73.0 vs 63.0; IQR, 59.0-67.0; P = .03) and on the mini-CEX (3.95; IQR, 3.63-4.20 vs 3.69; IQR, 3.36-3.90; P = .002), and they completed a greater percentage of required evaluations (95.6%; IQR, 88.1%-99.0% vs 86.1%; IQR, 70.6%-95.0%; P < .001) compared with residents with lower professionalism scores. In multivariate analysis, a professionalism score in the top 20% of residents was independently associated with ITE scores (odds ratio [OR] per 1-point increase, 1.07; 95% confidence interval [CI], 1.01-1.14; P = .046), mini-CEX scores (OR, 4.64; 95% CI, 1.23-17.48; P = .02), and completion of evaluations (OR, 1.07; 95% CI, 1.01-1.13; P = .02). Six of the 8 residents who received a warning or probation had total professionalism scores in the bottom 20% of residents. Observation-based assessments of professionalism were associated with residents' knowledge, clinical skills, and conscientious behaviors.