Background: Studies measuring physical activity (PA) and sedentary behavior on a 24/7 basis are scarce. The present study assessed the feasibility of using an accelerometer at the hip while awake and at the wrist while sleeping to describe 24/7 patterns of physical behavior in working-aged adults by age, sex, and fitness. Methods: The study was based on the FinFit 2017 study where the physical behavior of 20- to 69-year-old Finns was assessed 24/7 by triaxial accelerometer (UKKRM42; UKK Terveyspalvelut Oy, Tampere, Finland). During waking hours, the accelerometer was kept at the right hip and, during time in bed, at the nondominant wrist. PA variables were based on 1-min exponential moving average of mean amplitude deviation of the resultant acceleration signal analyzed in 6-s epochs. The angle for the posture estimation algorithm was used to identify sedentary behavior and standing. Evaluation of time in bed was based on the wrist movement. Fitness was estimated by the 6-min walk test. Results: A total of 2,256 eligible participants (mean age 49.5 years, SD = 13.5, 59% women) wore the accelerometer at the hip 15.7 hr/day (SD = 1.4) and at the wrist 8.3 hr/day (SD = 1.4). Sedentary behavior covered 9 hr 18 min/day (SD = 1.8 hr/day), standing nearly 2 hr/day (SD = 0.9), light PA 3.7 hr/day (SD = 1.3), and moderate to vigorous PA 46 min/day (SD = 26). Participants took 7,451 steps per day (SD = 2,962) on average. Men were most active around noon, while women had activity peaks at noon and at early evening. The low-fit tertile took 1,186 and 1,747 fewer steps per day than the mid- and high-fit tertiles (both p < .001). Conclusions: One triaxial accelerometer with a two wear-site approach provides a feasible method to characterize hour-by-hour patterns of physical behavior among working-aged adults.