Glycan Labeling and Analysis in Cells and In Vivo

As one of the major types of biomacromolecules in the cell, glycans play essential functional roles in various biological processes. Compared with proteins and nucleic acids, the analysis of glycans in situ has been more challenging. Herein we review recent advances in the development of methods and strategies for labeling, imaging, and profiling of glycans in cells and in vivo. Cellular glycans can be labeled by affinity-based probes, including lectin and antibody conjugates, direct chemical modification, metabolic glycan labeling, and chemoenzymatic labeling. These methods have been applied to label glycans with fluorophores, which enables the visualization and tracking of glycans in cells, tissues, and living organisms. Alternatively, labeling glycans with affinity tags has enabled the enrichment of glycoproteins for glycoproteomic profiling. Built on the glycan labeling methods, strategies enabling cell-selective and tissue-specific glycan labeling and protein-specific glycan imaging have been developed. With these methods and strategies, researchers are now better poised than ever to dissect the biological function of glycans in physiological or pathological contexts.

Real-Time Visualization and Monitoring of Physiological Dynamics by Aggregation-Induced Emission Luminogens (AIEgens)

Physiological dynamics in living cells and tissues are crucial for maintenance and regulation of their normal activities and functionalities. Tiny fluctuations in physiological microenvironments can leverage significant influences on cell growth, metabolism, differentiation, and apoptosis as well as disease evolution. Fluorescence imaging based on aggregation-induced emission luminogens (AIEgens) exhibits superior advantages in real-time sensing and monitoring of the physiological dynamics in living systems, including its unique properties such as high sensitivity and rapid response, flexible molecular design, and versatile nano- to mesostructural fabrication. The introduction of canonic AIEgens with long-wavelength, near-infrared, or microwave emission, persistent luminescence, and diversified excitation source (e.g., chemo- or bioluminescence) offers researchers a tool to evaluate the resulting molecules with excellent performance in response to subtle fluctuations in bioactivities with broader dimensionalities and deeper hierarchies.

Electrochemical Affinity Assays/Sensors: Brief History and Current Status

The advent of electrochemical affinity assays and sensors evolved from pioneering efforts in the 1970s to broaden the field of analytes accessible to the selective and sensitive performance of electrochemical detection. The foundation of electrochemical affinity assays/sensors is the specific capture of an analyte by an affinity element and the subsequent transduction of this event into a measurable signal. This review briefly covers the early development of affinity assays and then focuses on advances in the past decade. During this time, progress on electroactive labels, including the use of nanoparticles, quantum dots, organic and organometallic redox compounds, and enzymes with amplification schemes, has led to significant improvements in sensitivity. The emergence of nanomaterials along with microfabrication and microfluidics technology enabled research pathways that couple the ease of use of electrochemical detection for the development of devices that are more user friendly, disposable, and employable, such as lab-on-a-chip, paper, and wearable sensors.

Current Challenges and Recent Developments in Mass Spectrometry–Based Metabolomics

High-resolution mass spectrometry (MS) has advanced the study of metabolism in living systems by allowing many metabolites to be measured in a single experiment. Although improvements in mass detector sensitivity have facilitated the detection of greater numbers of analytes, compound identification strategies, feature reduction software, and data sharing have not kept up with the influx of MS data. Here, we discuss the ongoing challenges with MS-based metabolomics, including de novo metabolite identification from mass spectra, differentiation of metabolites from environmental contamination, chromatographic separation of isomers, and incomplete MS databases. Because of their popularity and sensitive detection of small molecules, this review focuses on the challenges of liquid chromatography-mass spectrometry–based methods. We then highlight important instrumentational, experimental, and computational tools that have been created to address these challenges and how they have enabled the advancement of metabolomics research.

Noncontact Nanoscale Imaging of Cells

The reduction in ion current as a fine pipette approaches a cell surface allows the cell surface topography to be imaged, with nanoscale resolution, without contact with the delicate cell surface. A variety of different methods have been developed and refined to scan the topography of the dynamic cell surface at high resolution and speed. Measurement of cell topography can be complemented by performing local probing or mapping of the cell surface using the same pipette. This can be done by performing single-channel recording, applying force, delivering agonists, using pipettes fabricated to contain an electrochemical probe, or combining with fluorescence imaging. These methods in combination have great potential to image and map the surface of live cells at the nanoscale.

Protein Dynamics by Two-Dimensional Infrared Spectroscopy

Proteins function as ensembles of interconverting structures. The motions span from picosecond bond rotations to millisecond and longer subunit displacements. Characterization of functional dynamics on all spatial and temporal scales remains challenging experimentally. Two-dimensional infrared spectroscopy (2D IR) is maturing as a powerful approach for investigating proteins and their dynamics. We outline the advantages of IR spectroscopy, describe 2D IR and the information it provides, and introduce vibrational groups for protein analysis. We highlight example studies that illustrate the power and versatility of 2D IR for characterizing protein dynamics and conclude with a brief discussion of the outlook for biomolecular 2D IR.

Recent Progress in the Analytical Chemistry of Champagne and Sparkling Wines

The strong interplay between the various parameters at play in a bottle and in a glass of champagne or sparkling wine has been the subject of study for about two decades. After a brief overview of the history of champagne and sparkling wines, this article presents the key steps involved in the traditional method leading to the production of premium modern-day sparkling wines, with a specific focus on quantification of the dissolved CO2 found in the sealed bottles. Moreover, a review of the literature on the various chemical and instrumental approaches used in the analysis of dissolved and gaseous CO2, effervescence, foam, and volatile organic compounds is reported. Expected final online publication date for the Annual Review of Analytical Chemistry, Volume 14 is August 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Developments and Ongoing Challenges for Analysis of Surface-Bound Proteins

Proteins at surfaces and interfaces play important roles in the function and performance of materials in applications ranging from diagnostic assays to biomedical devices. To improve the performance of these materials, detailed molecular structure (conformation and orientation) along with the identity and concentrations of the surface-bound proteins on those materials must be determined. This article describes radiolabeling, surface plasmon resonance, quartz crystal microbalance with dissipation, X-ray photoelectron spectroscopy, secondary ion mass spectrometry, sum frequency generation spectroscopy, and computational techniques along with the information each technique provides for characterizing protein films. A multitechnique approach using both experimental and computation methods is required for these investigations. Although it is now possible to gain much insight into the structure of surface-bound proteins, it is still not possible to obtain the same level of structural detail about proteins on surfaces as can be obtained about proteins in crystals and solutions, especially for large, complex proteins. However, recent results have shown it is possible to obtain detailed structural information (e.g., backbone and side chain orientation) about small peptides (5–20 amino sequences) on surfaces. Current studies are extending these investigations to small proteins such as protein G B1 (∼6 kDa). Approaches for furthering the capabilities for characterizing the molecular structure of surface-bound proteins are proposed. Expected final online publication date for the Annual Review of Analytical Chemistry, Volume 14 is August 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Active Flow Control and Dynamic Analysis in Droplet Microfluidics

Droplet-based microfluidics has emerged as an important subfield within the microfluidic and general analytical communities. Indeed, several unique applications such as digital assay readout and single-cell sequencing now have commercial systems based on droplet microfluidics. Yet there remains room for this research area to grow. To date, most analytical readouts are optical in nature, relatively few studies have integrated sample preparation, and passive means for droplet formation and manipulation have dominated the field. Analytical scientists continue to expand capabilities by developing droplet-compatible method adaptations, for example, by interfacing to mass spectrometers or automating droplet sampling for temporally resolved analysis. In this review, we highlight recently developed fluidic control techniques and unique integrations of analytical methodology with droplet microfluidics—focusing on automation and the connections to analog/digital domains—and we conclude by offering a perspective on current challenges and future applications. Expected final online publication date for the Annual Review of Analytical Chemistry, Volume 14 is August 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Clinical Chemistry for Developing Countries: Mass Spectrometry

Early disease diagnosis is necessary to enable timely interventions. Implementation of this vital task in the developing world is challenging owing to limited resources. Diagnostic approaches developed for resource-limited settings have often involved colorimetric tests (based on immunoassays) due to their low cost. Unfortunately, the performance/sensitivity of such simplistic tests are often limited and significantly hinder opportunities for early disease detection. A new criterion for selecting diagnostic tests in low- and middle-income countries is proposed here that is based on performance-to-cost ratio. For example, modern mass spectrometry (MS) now involves analysis of the native sample in the open laboratory environment, enabling applications in many fields, including clinical research, forensic science, environmental analysis, and agriculture. In this critical review, we summarize recent developments in chemistry that enable MS to be applied effectively in developing countries. In particular, we argue that closed automated analytical systems may not offer the analytical flexibility needed in resource-limited settings. Alternative strategies proposed here have potential to be widely accepted in low- and middle-income countries through the utilization of the open-source ambient MS platform that enables microsampling techniques such as dried blood spot to be coupled with miniature mass spectrometers in a centralized analytical platform. Consequently, costs associated with sample handling and maintenance can be reduced by >50% of the total ownership cost, permitting analytical measurements to be operated at high performance-to-cost ratios in the developing world. Expected final online publication date for the Annual Review of Analytical Chemistry, Volume 14 is August 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.