Visualization of microscale particle focusing in diluted and whole blood using particle trajectory analysis

Inertial microfluidics has demonstrated the potential to provide a rich range of capabilities to manipulate biological fluids and particles to address various challenges in biomedical science and clinical medicine. Various microchannel geometries have been used to study the inertial focusing behavior of particles suspended in simple buffer solutions or in highly diluted blood. One aspect of inertial focusing that has not been studied is how particles suspended in whole or minimally diluted blood respond to inertial forces in microchannels. The utility of imaging techniques (i.e., high-speed bright-field imaging and long exposure fluorescence (streak) imaging) primarily used to observe particle focusing in microchannels is limited in complex fluids such as whole blood due to interference from the large numbers of red blood cells (RBCs). In this study, we used particle trajectory analysis (PTA) to observe the inertial focusing behavior of polystyrene beads, white blood cells, and PC-3 prostate cancer cells in physiological saline and blood. Identification of in-focus (fluorescently labeled) particles was achieved at mean particle velocities of up to 1.85 m s(-1). Quantitative measurements of in-focus particles were used to construct intensity maps of particle frequency in the channel cross-section and scatter plots of particle centroid coordinates vs. particle diameter. PC-3 cells spiked into whole blood (HCT = 45%) demonstrated a novel focusing mode not observed in physiological saline or diluted blood. PTA can be used as an experimental frame of reference for understanding the physical basis of inertial lift forces in whole blood and discover inertial focusing modes that can be used to enable particle separation in whole blood.