School-based physical activity (PA), including activity during physical education (PE) lessons and after-school hours, is a possible opportunity for increasing children' s daily PA. The purposes of this study were (a) to compare children' s school-based PA levels during PE lessons and after-school hours and (b) to examine whether there is a difference in children' s PA behaviour (moderate-to-vigorous physical activity (MVPA)) and sedentary behaviour on a day which includes school-based PA (i.e. PE class or after-school sports class). The participants were 242 children (mean age = 8.7, SD = 1.6) from five primary schools in Hong Kong. Children' s PA levels were measured using an accelerometer (ActiGraph GT3X) for four school days, and the data were used to derive the duration (min) of MVPA and sedentary behaviour using age-specific criteria. The PA data were grouped according to the school-based PA opportunities, i.e. (a) with a PE lesson (PE day); (b) with an after-school sport class (AFS day); or (c) without school-based PA (N-Act day). The results indicated that the groups differed significantly in daily MVPA [F (2,108) = 16.62, p < .00] and MVPA during school hours [F (2,108) = 36.22, p < .00]. There was no significant difference in children' s sedentary behaviour with school-based PA participation. The present study confirmed the contribution of school-based PA opportunities to children' s MVPA in the school day, while sedentary behaviour may not necessarily be reduced.[Copyright © 2017 The Author(s).]