Label-free image-encoded microfluidic cell sorter with a scanning Bessel beam

Abstract

The microfluidic-based, label-free image-guided cell sorter offers a low-cost, high information content, and disposable solution that overcomes many limitations in conventional cell sorters. However, flow confinement for most microfluidic devices is generally only one-dimensional using sheath flow. As a result, the equilibrium distribution of cells spreads beyond the focal plane of commonly used Gaussian laser excitation beams, resulting in a large number of blurred images that hinder subsequent cell sorting based on cell image features. To address this issue, we present a Bessel–Gaussian beam image-guided cell sorter with an ultra-long depth of focus, enabling focused images of >85% of passing cells. This system features label-free sorting capabilities based on features extracted from the output temporal waveform of a photomultiplier tube (PMT) detector. For the sorting of polystyrene beads, SKNO1 leukemia cells, and Scenedesmus green algae, our results indicate a sorting purity of 97%, 97%, and 98%, respectively, showing that the temporal waveforms from the PMT outputs have strong correlations with cell image features. These correlations are also confirmed by off-line reconstructed cell images from a temporal–spatial transformation algorithm tailored to the scanning Bessel–Gaussian beam.

A high-throughput technique to map cell images to cell positions using a 3D imaging flow cytometer

“…The recent advances in the image-guided cell sorter or image-guided fluorescence-activated cell sorting (FACS) system have made good strides toward this goal ( 21 – 24 ). By isolating cells of the same image features based on a predefined gating criterion, one can perform a downstream genomic analysis with cells of similar imaging characteristics ( 21 , 22 , 24 , 25 ). However, today’s image-guided FACS produces only two-dimensional (2D) images, lacking the high information contents of three-dimensional (3D) imaging modalities such as confocal microscopes and light-sheet microscopes.…”