Flow Cytometry with Anti-Diffraction Light Sheet (ADLS) by Spatial Light Modulation

Abstract

Flow cytometry is a widespread and powerful technique whose resolution is determined by its capacity to accurately distinguish fluorescently positive populations from negative ones. However, most informative results are discarded while performing the measurements of conventional flow cytometry, e.g., the cell size, shape, morphology, and distribution or location of labeled exosomes within the unpurified biological samples. Herein, we propose a novel approach using an anti-diffraction light sheet with anisotroic feature to excite fluorescent tags. Constituted by an anti-diffraction Bessel–Gaussian beam array, the light sheet is 12 μm wide, 12 μm high, and has a thickness of ~0.8 μm. The intensity profile of the excited fluorescent signal can, therefore, reflect the size and allow samples in the range from O (100 nm) to 10 μm (e.g., blood cells) to be transported via hydrodynamic focusing in a microfluidic chip. The sampling rate is 500 kHz, which provides a capability of high throughput without sacrificing the spatial resolution. Consequently, the proposed anti-diffraction light sheet flow cytometry (ADLSFC) can obtain more informative results than the conventional methodologies, and is able to provide multiple characteristics (e.g., the size and distribution of fluorescent signal) helping to distinguish the target samples from the complex backgrounds.