The flow in a T-shaped micromixer at Reynolds numbers larger than Re = 100 becomes three-dimensional and intensive mixing occurs. To investigate the laminar, stationary three-dimensional flow in the T-mixer a stereo-μ-PIV system for the simultaneous measurement of all three components of the velocity vector field in a measurement plane (2D-3C) in a closed microchannel has been developed. Due to the very small confinement, standard calibration procedure by means of a calibration target is not possible, and therefore stereo-μ-PIV measurements in closed microchannels require a calibration based on the self-calibration of the tracer particle images. In order to include the effects of different refractive indices (of the fluid in the microchannel, the entrance window and the surrounding air) a three-media-model is included in the triangulation procedure of the self-calibration. Measurements in the mixing zone of a T-shaped micromixer at Re = 120 show that three-dimensional flow in a microchannel with dimensions of 800 × 200 μm can be measured with a resolution of 44 × 44 × 15 μm. The stationary flow in the 200 μm deep channel has been scanned in 22 μm wide steps, providing in a full 3D measurement of the averaged velocity distribution in the mixing zone of the T-mixer. The detailed full 3D measurement of the flow in the mixing region shows that the two inflows merge and bend over 90° in the clockwise/counter clockwise direction. When merging, the two inflows interact. The distribution of the liquid originating from the two inflows can be explained with the three-dimensional motion in the mixing zone. The liquid coming from the left inflow is deflected upwards, while the liquid coming from the right inflow is deflected downwards. Further downstream the two streams role up, causing the inflow from the right to move upwards on the lift side of the channel and vice versa for the inflow from the left. The complex three-dimensional flow structure increases the surface area between the two inflows, which leads to an increase of diffusive mixing of the laminar flow. In the region, where the two streams merge, the largest measured velocity gradient in the z direction is ∂νinplane/∂z = 2 · 104 l/s.

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