Dynamic pulse experiments for cell analysis require rapid and precise preparation of probes, which is often not possible in a macro-laboratory environment. Lab-on-a-Chip technology can offer new ways of probe preparation for both chemical and biochemical processes. A passive microfluidic mixer (micromixer) is presented in this contribution, which is designed for the preparation of cells. The micromixer is based on the method of split-and-recombination. Two alternating channel layers result in a three-dimensional pathway. Mixing in the laminar flow regime not only relies on molecular diffusion but is also enhanced by chaotic advection. The mixer was fabricated in a glass-silicon-glass sandwich technology, and mixing was characterized by chemical and biological probes. The contribution of chaotic advection, which appears in repeated 90° turns of the channel geometry, could be confirmed in computational fluid dynamics (CFD) analysis and laser-induced fluorescence images. Mixing performance was characterized by chemical iodometry. This method is based on the chemical reaction of Lugol’s solution and sodium thiosulfate. The resulting solution changes its color such that mixing becomes visible in a fluidic channel. Experiments were conducted for flow rates between 20 µL/min and 1000 µL/min corresponding to Reynolds numbers from 0.9 to 62. The experiments showed that fewer mixer units are necessary at higher flow rates because vorticity increases at higher Re in the recombination regions of the mixer. A mixing time of approximately 5 ms was achieved at a flow rate of 1000 µL/min at both inlets, which corresponds to a Re of 62 in this channel geometry. Biological pulse experiments were performed with the mixer, showing its suitability for preparing biological particles as eukaryotic cells.