A primary barrier to elucidating the association between sedentary behavior (SB) and health outcomes is the lack of valid monitors to assess SB in a free-living environment. The purpose of this study was to examine the validity of commercially available monitors to assess SB. Twenty overweight (mean ± SD: body mass index = 33.7 ± 5.7 kg·m(-2)) inactive, office workers age 46.5 ± 10.7 yr were directly observed for two 6-h periods while wearing an activPAL (AP) and an ActiGraph GT3X (AG). During the second observation, participants were instructed to reduce sitting time. We assessed the validity of the commonly used cut point of 100 counts per minute (AG100) and several additional AG cut points for defining SB. We used direct observation (DO) using focal sampling with duration coding to record either sedentary (sitting/lying) or nonsedentary behavior. The accuracy and precision of the monitors and the sensitivity of the monitors to detect reductions in sitting time were assessed using mixed-model repeated-measures analyses. On average, the AP and the AG100 underestimated sitting time by 2.8% and 4.9%, respectively. The correlation between the AP and DO was R2 = 0.94, and the AG100 and DO sedentary minutes was R2 = 0.39. Only the AP was able to detect reductions in sitting time. The AG 150-counts-per-minute threshold demonstrated the lowest bias (1.8%) of the AG cut points. The AP was more precise and more sensitive to reductions in sitting time than the AG, and thus, studies designed to assess SB should consider using the AP. When the AG monitor is used, 150 counts per minute may be the most appropriate cut point to define SB.