Abstract. We introduce a novel algorithm for actin filament segmentation in a 2D TIRFM image sequence. We treat the 2D time-lapse sequence as a 3D image volume and propose an over-grown active surface model to segment the body of a filament on all slices simultaneously. In order to locate the two ends of the filament on the over-grown surface, a novel 2D spatiotemporal domain is created based on the resulting surface. Two 2D active contour models deform in this domain to locate the two filament ends accurately. Evaluation on TIRFM image sequences with very low SNRs and comparison with a previous method demonstrate the accuracy and robustness of this approach.