Multi-ATOM: Ultrahigh-throughput single-cell quantitative phase imaging with subcellular resolution.

A growing body of evidence has substantiated the significance of quantitative phase imaging (QPI) in enabling cost-effective and label-free cellular assays, which provides useful insights into understanding the biophysical properties of cells and their roles in cellular functions. However, available QPI modalities are limited by the loss of imaging resolution at high throughput and thus run short of sufficient statistical power at the single-cell precision to define cell identities in a large and heterogeneous population of cells-hindering their utility in mainstream biomedicine and biology. Here we present a new QPI modality, coined multiplexed asymmetric-detection time-stretch optical microscopy (multi-ATOM) that captures and processes quantitative label-free single-cell images at ultrahigh throughput without compromising subcellular resolution.

Multi-MHz laser-scanning single-cell fluorescence microscopy by spatiotemporally encoded virtual source array.

Apart from the spatial resolution enhancement, scaling of temporal resolution, equivalently the imaging throughput, of fluorescence microscopy is of equal importance in advancing cell biology and clinical diagnostics. Yet, this attribute has mostly been overlooked because of the inherent speed limitation of existing imaging strategies. To address the challenge, we employ an all-optical laser-scanning mechanism, enabled by an array of reconfigurable spatiotemporally-encoded virtual sources, to demonstrate ultrafast fluorescence microscopy at line-scan rate as high as 8 MHz.

Microfluidic Imaging Flow Cytometry by Asymmetric-detection Time-stretch Optical Microscopy (ATOM).

Scaling the number of measurable parameters, which allows for multidimensional data analysis and thus higher-confidence statistical results, has been the main trend in the advanced development of flow cytometry. Notably, adding high-resolution imaging capabilities allows for the complex morphological analysis of cellular/sub-cellular structures. This is not possible with standard flow cytometers.

Time-stretch microscopy on a DVD for high-throughput imaging cell-based assay.

Cell-based assay based on time-stretch imaging is recognized to be well-suited for high-throughput phenotypic screening. However, this ultrafast imaging technique has primarily been limited to suspension-cell assay, leaving a wide range of solid-substrate assay formats uncharted. Moreover, time-stretch imaging is generally restricted to intrinsic biophysical phenotyping, but lacks the biomolecular signatures of the cells.

High-throughput time-stretch imaging flow cytometry for multi-class classification of phytoplankton.

Time-stretch imaging has been regarded as an attractive technique for high-throughput imaging flow cytometry primarily owing to its real-time, continuous ultrafast operation. Nevertheless, two key challenges remain: (1) sufficiently high time-stretch image resolution and contrast is needed for visualizing sub-cellular complexity of single cells, and (2) the ability to unravel the heterogeneity and complexity of the highly diverse population of cells - a central problem of single-cell analysis in life sciences - is required. We here demonstrate an optofluidic time-stretch imaging flow cytometer that enables these two features, in the context of high-throughput multi-class (up to 14 classes) phytoplantkton screening and classification.

Asymmetric-detection time-stretch optical microscopy (ATOM) for ultrafast high-contrast cellular imaging in flow.

Accelerating imaging speed in optical microscopy is often realized at the expense of image contrast, image resolution, and detection sensitivity--a common predicament for advancing high-speed and high-throughput cellular imaging. We here demonstrate a new imaging approach, called asymmetric-detection time-stretch optical microscopy (ATOM), which can deliver ultrafast label-free high-contrast flow imaging with well delineated cellular morphological resolution and in-line optical image amplification to overcome the compromised imaging sensitivity at high speed. We show that ATOM can separately reveal the enhanced phase-gradient and absorption contrast in microfluidic live-cell imaging at a flow speed as high as ~10 m/s, corresponding to an imaging throughput of ~100,000 cells/sec.