Photoswitchable polyynes for multiplexed stimulated Raman scattering microscopy with reversible light control.

Optical imaging with photo-controllable probes has greatly advanced biological research. With superb chemical specificity of vibrational spectroscopy, stimulated Raman scattering (SRS) microscopy is particularly promising for super-multiplexed optical imaging with rich chemical information. Functional SRS imaging in response to light has been recently demonstrated, but multiplexed SRS imaging with reversible photocontrol remains unaccomplished.

Plasmon-Induced Charge Transfer-Enhanced Raman Scattering on a Semiconductor: Toward Amplification-Free Quantification of SARS-CoV-2.

Semiconductors demonstrate great potentials as chemical mechanism-based surface-enhanced Raman scattering (SERS) substrates in determination of biological species in complex living systems with high selectivity. However, low sensitivity is the bottleneck for their practical applications, compared with that of noble metal-based Raman enhancement ascribed to electromagnetic mechanism. Herein, a novel Cu O nanoarray with free carrier density of 1.

Identification of essential sites of lipid peroxidation in ferroptosis.

Ferroptosis, an iron-dependent form of cell death driven by lipid peroxidation, provides a potential treatment avenue for drug-resistant cancers and may play a role in the pathology of some degenerative diseases. Identifying the subcellular membranes essential for ferroptosis and the sequence of their peroxidation will illuminate drug discovery strategies and ferroptosis-relevant disease mechanisms. In this study, we employed fluorescence and stimulated Raman scattering imaging to examine the structure-activity-distribution relationship of ferroptosis-modulating compounds.

Super-multiplex imaging of cellular dynamics and heterogeneity by integrated stimulated Raman and fluorescence microscopy.

Observing multiple molecular species simultaneously with high spatiotemporal resolution is crucial for comprehensive understanding of complex, dynamic, and heterogeneous biological systems. The recently reported super-multiplex optical imaging breaks the "color barrier" of fluorescence to achieve multiplexing number over six in living systems, while its temporal resolution is limited to several minutes mainly by slow color tuning. Herein, we report integrated stimulated Raman and fluorescence microscopy with simultaneous multimodal color tunability at high speed, enabling super-multiplex imaging covering diverse molecular contrasts with temporal resolution of seconds.

9-Cyanopyronin probe palette for super-multiplexed vibrational imaging.

Multiplexed optical imaging provides holistic visualization on a vast number of molecular targets, which has become increasingly essential for understanding complex biological processes and interactions. Vibrational microscopy has great potential owing to the sharp linewidth of vibrational spectra. In 2017, we demonstrated the coupling between electronic pre-resonant stimulated Raman scattering (epr-SRS) microscopy with a proposed library of 9-cyanopyronin-based dyes, named Manhattan Raman Scattering (MARS).

Ultra-bright Raman dots for multiplexed optical imaging.

Imaging the spatial distribution of biomolecules is at the core of modern biology. The development of fluorescence techniques has enabled researchers to investigate subcellular structures with nanometer precision. However, multiplexed imaging, i.

Optical mapping of biological water in single live cells by stimulated Raman excited fluorescence microscopy.

Water is arguably the most common and yet least understood material on Earth. Indeed, the biophysical behavior of water in crowded intracellular milieu is a long-debated issue. Understanding of the spatial and compositional heterogeneity of water inside cells remains elusive, largely due to a lack of proper water-sensing tools with high sensitivity and spatial resolution.

Biological imaging of chemical bonds by stimulated Raman scattering microscopy.

All molecules consist of chemical bonds, and much can be learned from mapping the spatiotemporal dynamics of these bonds. Since its invention a decade ago, stimulated Raman scattering (SRS) microscopy has become a powerful modality for imaging chemical bonds with high sensitivity, resolution, speed and specificity. We introduce the fundamentals of SRS microscopy and review innovations in SRS microscopes and imaging probes.

Structure-activity-distribution relationship study of anti-cancer antimycin-type depsipeptides.

Small-molecule natural products have been an essential source of pharmaceuticals to treat human diseases, but very little is known about their behavior inside dynamic, live human cells. Here, we demonstrate the first structure-activity-distribution relationship (SADR) study of complex natural products, the anti-cancer antimycin-type depsipeptides, using the emerging bioorthogonal Stimulated Raman Scattering (SRS) Microscopy. Our results show that the intracellular enrichment and distribution of these compounds are driven by their potency and specific protein targets, as well as the lipophilic nature of compounds.

Raman Imaging of Small Biomolecules.

Imaging techniques greatly facilitate the comprehensive knowledge of biological systems. Although imaging methodology for biomacromolecules such as protein and nucleic acids has been long established, microscopic techniques and contrast mechanisms are relatively limited for small biomolecules, which are equally important participants in biological processes. Recent developments in Raman imaging, including both microscopy and tailored vibrational tags, have created exciting opportunities for noninvasive imaging of small biomolecules in living cells, tissues, and organisms.

Probe design for super-multiplexed vibrational imaging.

Optical microscopy has served biomedical research for decades due to its high temporal and spatial resolutions. Among various optical imaging techniques, fluorescence imaging offers superb sensitivity down to single molecule level but its multiplexing capacity is limited by intrinsically broad bandwidth. To simultaneously capture a vast number of targets, the newly emerging vibrational microscopy technique draws increasing attention as vibration spectroscopy features narrow transition linewidth.

A ratiometric Raman probe for live-cell imaging of hydrogen sulfide in mitochondria by stimulated Raman scattering.

Stimulated Raman Scattering (SRS) coupled with alkyne tags has been an emerging imaging technique to visualize small-molecule species with high sensitivity and specificity. Here we describe the development of a ratiometric Raman probe for visualizing hydrogen sulfide (H2S) species in living cells as the first alkyne-based sensor for SRS microscopy. This probe uses an azide unit as a selective reactive site, and it targets mitochondria with high specificity.

Determination of the Subcellular Localization and Mechanism of Action of Ferrostatins in Suppressing Ferroptosis.

Ferroptosis is a form of nonapoptotic cell death characterized by the unchecked accumulation of lipid peroxides. Ferrostatin-1 and its analogs (ferrostatins) specifically prevent ferroptosis in multiple contexts, but many aspects of their molecular mechanism of action remain poorly described. Here, we employed stimulated Raman scattering (SRS) microscopy coupled with small vibrational tags to image the distribution of ferrostatins in cells and found that they accumulate in lysosomes, mitochondria, and the endoplasmic reticulum.

Supermultiplexed optical imaging and barcoding with engineered polyynes.

Optical multiplexing has a large impact in photonics, the life sciences and biomedicine. However, current technology is limited by a 'multiplexing ceiling' from existing optical materials. Here we engineered a class of polyyne-based materials for optical supermultiplexing.

Two-color vibrational imaging of glucose metabolism using stimulated Raman scattering.

A two-color vibrational imaging technique for simultaneously mapping glucose uptake and incorporation activity inside single living cells is reported. Heterogeneous patterns of glucose metabolism are directly visualized from the ratiometric two-color images for various cell types, cells undergoing epithelia-to-mesenchymal transitions and live mouse brain tissues. The two-color imaging of glucose metabolism here demonstrates the development of multi-functional vibrational probes for multicolor imaging of cellular metabolism.

Applications of vibrational tags in biological imaging by Raman microscopy.

As a superb tool to visualize and study the spatial-temporal distribution of chemicals, Raman microscopy has made a big impact in many disciplines of science. While label-free imaging has been the prevailing strategy in Raman microscopy, recent development and applications of vibrational/Raman tags, particularly when coupled with stimulated Raman scattering (SRS) microscopy, have generated intense excitement in biomedical imaging. SRS imaging of vibrational tags has enabled researchers to study a wide range of small biomolecules with high specificity, sensitivity and multiplex capability, at a single live cell level, tissue level or even in vivo.

Stimulated Raman scattering of polymer nanoparticles for multiplexed live-cell imaging.

A novel nanoparticle-based imaging strategy is introduced that couples biocompatible organic nanoparticles and stimulated Raman scattering (SRS) microscopy. Polymer nanoparticles with vibrational labels incorporated were readily prepared for multi-color SRS imaging with excellent photo-stability. The Raman-active polymer dots are nontoxic, rapidly enter various cell types, and are applied in multiplexed cell-type sorting.

Super-multiplex vibrational imaging.

The ability to visualize directly a large number of distinct molecular species inside cells is increasingly essential for understanding complex systems and processes. Even though existing methods have successfully been used to explore structure-function relationships in nervous systems, to profile RNA in situ, to reveal the heterogeneity of tumour microenvironments and to study dynamic macromolecular assembly, it remains challenging to image many species with high selectivity and sensitivity under biological conditions. For instance, fluorescence microscopy faces a 'colour barrier', owing to the intrinsically broad (about 1,500 inverse centimetres) and featureless nature of fluorescence spectra that limits the number of resolvable colours to two to five (or seven to nine if using complicated instrumentation and analysis).

Bioorthogonal chemical imaging of metabolic activities in live mammalian hippocampal tissues with stimulated Raman scattering.

Brain is an immensely complex system displaying dynamic and heterogeneous metabolic activities. Visualizing cellular metabolism of nucleic acids, proteins, and lipids in brain with chemical specificity has been a long-standing challenge. Recent development in metabolic labeling of small biomolecules allows the study of these metabolisms at the global level.

Macrosteres: The Deltic Guanidinium Ion.

The "deltic guanidinium" ion is described here as a "macrostere" of the guanidinium ion. The use of the 2,4-dimethoxybenzyl protecting group allows for the synthesis of the fully unsubstituted parent compound and a variety of derivatives bearing multiple N-H functions for the first time. Deltic urea, deltic thiourea, and deltic benzamidine are also synthesized.

Live-Cell Bioorthogonal Chemical Imaging: Stimulated Raman Scattering Microscopy of Vibrational Probes.

Innovations in light microscopy have tremendously revolutionized the way researchers study biological systems with subcellular resolution. In particular, fluorescence microscopy with the expanding choices of fluorescent probes has provided a comprehensive toolkit to tag and visualize various molecules of interest with exquisite specificity and high sensitivity. Although fluorescence microscopy is currently the method of choice for cellular imaging, it faces fundamental limitations for studying the vast number of small biomolecules.

Vibrational Imaging of Glucose Uptake Activity in Live Cells and Tissues by Stimulated Raman Scattering.

Glucose is a ubiquitous energy source for most living organisms. Its uptake activity closely reflects cellular metabolic demand in various physiopathological conditions. Extensive efforts have been made to specifically image glucose uptake, such as with positron emission tomography, magnetic resonance imaging, and fluorescence microscopy, but all have limitations.