Imaging human blood cells in vivo with oblique back-illumination capillaroscopy

Abstract: We present a non-invasive, label-free method of imaging blood cells flowing through human capillaries in vivo using oblique back-illumination capillaroscopy (OBC). Green light illumination allows simultaneous phase and absorption contrast, enhancing the ability to distinguish red and white blood cells. Single-sided illumination through the objective lens enables 200 Hz imaging with close illumination-detection separation and a simplified setup. Phase contrast is optimized when the illumination axis is offset from the detection axis by approximately 225 um when imaging 80 um deep in phantoms and human ventral tongue. We demonstrate high-speed imaging of individual red blood cells, white blood cells with sub-cellular detail, and platelets flowing through capillaries and vessels in human tongue. A custom pneumatic cap placed over the objective lens stabilizes the field of view, enabling longitudinal imaging of a single capillary for up to seven minutes. We present high-quality images of blood cells in individuals with Fitzpatrick skin phototypes II, IV, and VI, showing that the technique is robust to high peripheral melanin concentration. The signal quality, speed, simplicity, and robustness of this approach underscores its potential for non-invasive blood cell counting.