High-speed velocimetry in microfluidic protein mixers using confocal fluorescence decay microscopy

A method to measure fluid speeds on the order of 10,000 mm/s in microchannels is presented. A microfluidic protein mixer is manufactured with a 170 µm microscope coverslip bottom that interfaces with a confocal florescence microscope using a water-immersed Olympus UPLSAPO 60XW objective to create a diffraction-limited confocal volume. A diode laser with a repetition rate of 1 MHz is used to study Poiseuille flows at average speeds of 5000, 6000 and 7000 mm/s by exciting tris(2,2′-bipyridine)ruthenium(II) hexafluorophosphate solution at 1 μmol/L concentration flowing through the micro-mixer in the confocal volume. Decays collected using a time-correlated single photon counting card at each grid point are characterized by the first moment of the decay and curve fitted with the theoretical Poiseuille flow solutions. It was found that curve fitting with higher average speeds results in lower errors. A fluorescence correlation study was then carried out at different depths in the micro-mixer to understand the raw data profiles observed using the diode laser. A mixing study was then carried out using a Ti-Sapphire laser with a repetition rate of 3.8 MHz. A Poiseuille flow at 7000 mm/s was measured using the Ti-Sapphire laser and then curve fitted to the theoretical Poiseuille flow solution. The curve fit was then applied to the complicated flow region to determine speed. Results of the experimental mixing study are also compared to direct numerical simulation results. Graphical abstract An experimental technique to measure speeds in the complicated mixing region of a protein mixer is developed. Results show that when flow is bounded asymmetrically between a rough wall and smooth wall, the flow shows affinity for the smooth wall during the mixing process. The Reynolds number for this is flow is 245 indicating flow in the transitional turbulence regime.