Interferometric time-stretch microscopy for ultrafast quantitative cellular and tissue imaging at 1 μm

Quantitative phase imaging (QPI) has been proven to be a powerful tool for label-free characterization of biological specimens. However, the imaging speed, largely limited by the image sensor technology, impedes its utility in applications where high-throughput screening and efficient big-data analysis are mandated. We here demonstrate interferometric time-stretch (iTS) microscopy for delivering ultrafast quantitative phase cellular and tissue imaging at an imaging line-scan rate >20 MHz-orders-of-magnitude faster than conventional QPI. Enabling an efficient time-stretch operation in the 1-mum wavelength window, we present an iTS microscope system for practical ultrafast QPI of fixed cells and tissue sections, as well as ultrafast flowing cells (at a flow speed of up to 8 ms). To the best of our knowledge, this is the first time that time-stretch imaging could reveal quantitative morphological information of cells and tissues with nanometer precision. As many parameters can be further extracted from the phase and can serve as the intrinsic biomarkers for disease diagnosis, iTS microscopy could find its niche in high-throughput and high-content cellular assays (e.g., imaging flow cytometry) as well as tissue refractometric imaging (e.g., whole-slide imaging for digital pathology).published_or_final_versio

Ultrafast Laser-Scanning Time-Stretch Imaging at Visible Wavelengths

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

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Optofluidic time-stretch imaging – an emerging tool for high-throughput imaging flow cytometry

Optical imaging is arguably the most effective tool to visualize living cells with high spatiotemporal resolu- tion and in a nearly noninvasive manner. Driven by this capability, state-of-the-art cellular assay techniques have increasingly been adopting optical imaging for classifying different cell types/stages, and thus dissecting the respective cellular functions. However, it is still a daunting task to image and characterize cell-to-cell variability within an enormous and heterogeneous population – an unmet need in single-cell analysis, which is now widely advocated in modern biology and clinical diagnostics. The challenge stems from the fact that current optical imaging technologies still lack the practical speed and sensitivity for mea- suring thousands to millions of cells down to the single-cell precision. Adopting the wisdom in high-speed fiber-optics communication, optical time-stretch imaging has emerged as a completely new optical imag- ing concept which is now proven for ultrahigh-throughput optofluidic single-cell imaging, at least 1–2 orders-of-magnitude higher (up to ∼100000 cells per second) compared to the existing imaging flow cytometers. It also uniquely enables quantification of intrinsic biophysical markers of individual cells – a largely unexploited class of single-cell signatures that is known to be correlated with the overwhelmingly investigated biochemical markers. With the aim of reaching a wider spectrum of experts specializing in cel- lular assay developments and applications, this paper highlights the essential basics of optical time-stretch imaging, followed by reviewing the recent developments and applications of optofluidic time-stretch imag- ing. We will also discuss the current challenges of this technology, in terms of providing new insights in ba- sic biology and enriching the clinical diagnostic toolsets

Methods in Molecular Biology

There is an unmet need in biomedicine for measuring a multitude of parameters of individual cells (i.e., high content) in a large population ef ciently (i.e., high throughput). This is particularly driven by the emerging interest in bringing Big-Data analysis into this arena, encompassing pathology, drug discovery, rare cancer cell detection, emulsion microdroplet assays, to name a few. This momentum is particularly evident in recent advancements in ow cytometry. They include scaling of the number of measurable colors from the labeled cells and incorporation of imaging capability to access the morphological information of the cells. However, an unspoken predicament appears in the current technologies: higher content comes at the expense of lower throughput, and vice versa. For example, accessing additional spatial information of individual cells, imaging ow cytometers only achieve an imaging throughput ~1000 cells/s, orders of magnitude slower than the non- imaging ow cytometers. In this chapter, we introduce an entirely new imaging platform, namely optical time-stretch microscopy, for ultrahigh speed and high contrast label-free single-cell (in a ultrafast micro uidic ow up to 10 m/s) imaging and analysis with an ultra-fast imaging line-scan rate as high as tens of MHz. Based on this technique, not only morphological information of the individual cells can be obtained in an ultrafast manner, quantitative evaluation of cellular information (e.g., cell volume, mass, refractive index, stiffness, membrane tension) at nanometer scale based on the optical phase is also possible. The technology can also be integrated with conventional uorescence measurements widely adopted in the non-imaging ow cytometers. Therefore, these two combinatorial and complementary measurement capabilities in long run is an attractive platform for addressing the pressing need for expanding the “parameter space” in high-throughput single-cell analysis. This chapter provides the general guidelines of constructing the optical system for time stretch imaging, fabrication and design on the micro uidic chip for ultrafast uidic ow, as well as the image acquisition and processing

Optical Time Stretch for High-Speed and High-Throughput Imaging—From Single-Cell to Tissue-Wide Scales

Initially developed for high-speed optical communica- tion, optical time stretch has recently been adopted for ultrafast and sensitive optical imaging at an unprecedented speed. In this paper, we highlight the essential concepts as well as the enabling elements of this ultrafast technology. More importantly, we review the re- cent developments of optical time-stretch imaging, especially in the context of 1) quantitative optofluidic microscopy for high-content single-cell phenotyping at an imaging throughput ∼100 000 cells/s; 2) all-optical multi-MHz (>10 MHz) swept-source optical coher- ence tomography (OCT) for high-speed in vivo anatomical and functional 3-D tissue imaging. We also discuss the current techno- logical challenges in time-stretch imaging. In particular, generating the enormous data in real time, this technology could uniquely cre- ate new insights of data-driven science in clinical diagnostics and basic biological research

Quantitative asymmetric-detection time-stretch optical microscopy (Q-ATOM) for ultrafast quantitative phase imaging flow cytometry

Asymmetric-detection time-stretch optical microscopy (ATOM) has recently been developed for label-free high-contrast ultrafast single-cell imaging. We demonstrate that this technique can be further advanced, termed quantitative ATOM (Q-ATOM) to acquire quantitative-phase images of single cells at an ultrafast line-scan rate beyond 10 MHz (an equivalent imaging throughput of ~100,000 cells/sec) based on multiple off-axis light beam detection. Without the need for interferometry and computationally intensive phase-retrieval algorithms, Q-ATOM could be a robust approach for high-throughput label-free single-cell phenotyping based on their intrinsic biophysical markers