Fig. 3: Full 3D vector flow characterization and influence of sample orientation.

a Workflow to determine the 3D velocity field over the imaging volume from the measured tracer particle trajectories. First, the instantaneous velocity vectors, u, are extracted for every track. Next, the imaging volume is divided into voxels and all instantaneous velocity vectors within a given voxel are collected for subsequent analysis. Then, from the instantaneous velocity vector distribution of said voxel, an ensemble average flow velocity vector, U, is determined. This process suppresses the Brownian motion contribution of each tracer particle in U. Finally, this procedure is repeated for all the remaining voxels. b Representative 3D particle trajectories, with time evolution encoded in color, together with the resulting 3D velocity field map for an optofluidic platform with a nominal channel height of 50 µm oriented perpendicular to the direction of gravity as shown in the schematic diagram. c Same as (b) but for an optofluidic platform that is oriented parallel to the direction of gravity. The scheme indicates the plane along which the maps in (b) and (c) are calculated given the change in orientation of the optical axis, XY and XZ respectively. Each row and column in the velocity maps correspond to a different vector component and channel depth position, respectively. Color encodes the direction and magnitude of the flow velocity, whereas the solid lines represent a contour line indicating a change in direction. All velocity vector components share the same magnitude scale, which has been normalized by the maximum temperature increase in the system.