Background: Regular activity breaks positively impact markers of cardiometabolic health when performed in a laboratory. However, identifying compliance to a free-living regular activity breaks intervention is challenging, particularly if intensity is prescribed. Methods: This study had two parts. In Part A, 20 participants performed activity breaks similar to those shown to impart health benefits while wearing an ActiGraph and activPAL accelerometer, and a heart rate monitor. In Part B, the threshold found to identify these activities was used to identify the activity breaks performed by 78 sedentary, university employees wearing an ActiGraph accelerometer for seven days. Results: A cut-point of 1,000 vector magnitude counts per minute accurately identified activity breaks performed in the laboratory. Applying this cut-point to data collected in free living, sedentary participants identified, on average, seven activity breaks were being performed during work-hours. Conclusions: Using a cut-point of 1,000 vector magnitude counts per minute will identify activity breaks of a similar intensity to those found to elicit acute cardiometabolic benefit. Sedentary university employees may benefit from interventions to increase the number of activity breaks performed across their entire day.