Label-Free, Real-Time Measurement of Metabolism of Adherent and Suspended Single Cells by In-Cell Fourier Transform Infrared Microspectroscopy

We used infrared (IR) microscopy to monitor in real-time the metabolic turnover of individual mammalian cells in morphologically different states. By relying on the intrinsic absorption of mid-IR light by molecular components, we could discriminate the metabolism of adherent cells as compared to suspended cells. We identified major biochemical differences between the two cellular states, whereby only adherent cells appeared to rely heavily on glycolytic turnover and lactic fermentation. We also report spectroscopic variations that appear as spectral oscillations in the IR domain, observed only when using synchrotron infrared radiation. We propose that this effect could be used as a reporter of the cellular conditions. Our results are instrumental in establishing IR microscopy as a label-free method for real-time metabolic studies of individual cells in different morphological states, and in more complex cellular ensembles.

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Gating a Single Cell: A Label-Free and Real-Time Measurement Method for Cellular Progression

Analytical Chemistry ◽

10.1021/acs.analchem.9b03136 ◽

2020 ◽

Vol 92 (2) ◽

pp. 1738-1745

Author(s):

Saw Lin Oo ◽

Shishir Venkatesh ◽

Abdul-Mojeed Ilyas ◽

Vaithinathan Karthikeyan ◽

Clement Manohar Arava ◽

...

Keyword(s):

Single Cell ◽

Real Time ◽

Measurement Method ◽

Time Measurement ◽

Label Free ◽

Real Time Measurement

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Advanced Optogenetic-Based Biosensing and Related Biomaterials

Materials ◽

10.3390/ma14154151 ◽

2021 ◽

Vol 14 (15) ◽

pp. 4151

Author(s):

Mihaela Gheorghiu ◽

Cristina Polonschii ◽

Octavian Popescu ◽

Eugen Gheorghiu

Keyword(s):

Real Time ◽

Mammalian Cells ◽

Label Free ◽

Sensing Platforms ◽

Low Concentrations ◽

Cellular Diagnostics ◽

Cellular Sensors ◽

Higher Sensitivity ◽

Sensitivity Detection ◽

Optogenetic Control

The ability to stimulate mammalian cells with light, brought along by optogenetic control, has significantly broadened our understanding of electrically excitable tissues. Backed by advanced (bio)materials, it has recently paved the way towards novel biosensing concepts supporting bio-analytics applications transversal to the main biomedical stream. The advancements concerning enabling biomaterials and related novel biosensing concepts involving optogenetics are reviewed with particular focus on the use of engineered cells for cell-based sensing platforms and the available toolbox (from mere actuators and reporters to novel multifunctional opto-chemogenetic tools) for optogenetic-enabled real-time cellular diagnostics and biosensor development. The key advantages of these modified cell-based biosensors concern both significantly faster (minutes instead of hours) and higher sensitivity detection of low concentrations of bioactive/toxic analytes (below the threshold concentrations in classical cellular sensors) as well as improved standardization as warranted by unified analytic platforms. These novel multimodal functional electro-optical label-free assays are reviewed among the key elements for optogenetic-based biosensing standardization. This focused review is a potential guide for materials researchers interested in biosensing based on light-responsive biomaterials and related analytic tools.

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Real-time measurement of intracellular O2 in mammalian cells

BioTechniques ◽

10.2144/000114539 ◽

2017 ◽

Vol 62 (4) ◽

Keyword(s):

Real Time ◽

Mammalian Cells ◽

Time Measurement ◽

Real Time Measurement

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Advanced Biosensing towards Real-Time Imaging of Protein Secretion from Single Cells

10.5772/intechopen.94248 ◽

2020 ◽

Author(s):

Lang Zhou ◽

Pengyu Chen ◽

Aleksandr Simonian

Keyword(s):

Single Cell ◽

Real Time ◽

Protein Secretion ◽

Intercellular Communication ◽

Single Cells ◽

Bone Marrow Failure ◽

Cellular Protein ◽

Label Free ◽

Immune Effector ◽

Real Time Imaging

Protein secretion of cells plays a vital role in intercellular communication. The abnormality and dysfunction of cellular protein secretion are associated with various physiological disorders, such as malignant proliferation of cells, aberrant immune function, and bone marrow failure. The heterogeneity of protein secretion exists not only between varying populations of cells, but also in the same phenotype of cells. Therefore, characterization of protein secretion from single cell contributes not only to the understanding of intercellular communication in immune effector, carcinogenesis and metastasis, but also to the development and improvement of diagnosis and therapy of relative diseases. In spite of abundant highly sensitive methods that have been developed for the detection of secreted proteins, majority of them fall short in providing sufficient spatial and temporal resolution for comprehensive profiling of protein secretion from single cells. The real-time imaging techniques allow rapid acquisition and manipulation of analyte information on a 2D plane, providing high spatiotemporal resolution. Here, we summarize recent advances in real-time imaging of secretory proteins from single cell, including label-free and labelling techniques, shedding light on the development of simple yet powerful methodology for real-time imaging of single-cell protein secretion.

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All-digital histopathology by infrared-optical hybrid microscopy

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1912400117 ◽

2020 ◽

Vol 117 (7) ◽

pp. 3388-3396 ◽

Cited By ~ 8

Author(s):

Martin Schnell ◽

Shachi Mittal ◽

Kianoush Falahkheirkhah ◽

Anirudh Mittal ◽

Kevin Yeh ◽

...

Keyword(s):

Optical Microscopy ◽

Digital Pathology ◽

Cost Effective ◽

Optical Microscope ◽

Wide Field ◽

Label Free ◽

Ir Microscopy ◽

Almost All ◽

Molecular Components ◽

Ir Light

Optical microscopy for biomedical samples requires expertise in staining to visualize structure and composition. Midinfrared (mid-IR) spectroscopic imaging offers label-free molecular recording and virtual staining by probing fundamental vibrational modes of molecular components. This quantitative signal can be combined with machine learning to enable microscopy in diverse fields from cancer diagnoses to forensics. However, absorption of IR light by common optical imaging components makes mid-IR light incompatible with modern optical microscopy and almost all biomedical research and clinical workflows. Here we conceptualize an IR-optical hybrid (IR-OH) approach that sensitively measures molecular composition based on an optical microscope with wide-field interferometric detection of absorption-induced sample expansion. We demonstrate that IR-OH exceeds state-of-the-art IR microscopy in coverage (10-fold), spatial resolution (fourfold), and spectral consistency (by mitigating the effects of scattering). The combined impact of these advances allows full slide infrared absorption images of unstained breast tissue sections on a visible microscope platform. We further show that automated histopathologic segmentation and generation of computationally stained (stainless) images is possible, resolving morphological features in both color and spatial detail comparable to current pathology protocols but without stains or human interpretation. IR-OH is compatible with clinical and research pathology practice and could make for a cost-effective alternative to conventional stain-based protocols for stainless, all-digital pathology.

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Real-Time Monitoring of the Catalytic Oxidation of Alcohols to Aldehydes and Ketones on Resin Support by Single-Bead Fourier Transform Infrared Microspectroscopy

The Journal of Organic Chemistry ◽

10.1021/jo9613895 ◽

1996 ◽

Vol 61 (25) ◽

pp. 8765-8770 ◽

Cited By ~ 35

Author(s):

Bing Yan ◽

Qun Sun ◽

James R. Wareing ◽

Charles F. Jewell

Keyword(s):

Fourier Transform ◽

Real Time ◽

Catalytic Oxidation ◽

Fourier Transform Infrared ◽

Real Time Monitoring ◽

Infrared Microspectroscopy ◽

Fourier Transform Infrared Microspectroscopy ◽

Oxidation Of Alcohols ◽

Aldehydes And Ketones ◽

Single Bead

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Real-Time Monitoring and Detection of Single-Cell Level Cytokine Secretion Using LSPR Technology

Micromachines ◽

10.3390/mi11010107 ◽

2020 ◽

Vol 11 (1) ◽

pp. 107 ◽

Cited By ~ 3

Author(s):

Chen Zhu ◽

Xi Luo ◽

Wilfred Villariza Espulgar ◽

Shohei Koyama ◽

Atsushi Kumanogoh ◽

...

Keyword(s):

Single Cell ◽

Real Time ◽

Single Cells ◽

Clinical Diagnostics ◽

Cytokine Secretion ◽

Localized Surface Plasmon ◽

Single Cell Level ◽

Label Free ◽

Real Time Monitoring ◽

Cell Level

Cytokine secretion researches have been a main focus of studies among the scientists in the recent decades for its outstanding contribution to clinical diagnostics. Localized surface plasmon resonance (LSPR) technology is one of the conventional methods utilized to analyze these issues, as it could provide fast, label-free and real-time monitoring of biomolecule binding events. However, numerous LSPR-based biosensors in the past are usually utilized to monitor the average performance of cell groups rather than single cells. Meanwhile, the complicated sensor structures will lead to the fabrication and economic budget problems. Thus, in this paper, we report a simple synergistic integration of the cell trapping of microwell chip and gold-capped nanopillar-structured cyclo-olefin-polymer (COP) film for single cell level Interleukin 6 (IL-6) detection. Here, in-situ cytokine secreted from the trapped cell can be directly observed and analyzed through the peak red-shift in the transmittance spectrum. The fabricated device also shows the potential to conduct the real-time monitoring which would greatly help us identify the viability and biological variation of the tested single cell.

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Dissociation Constant of Integrin-RGD Binding in Live Cells from Automated Micropipette and Label-Free Optical Data

Biosensors ◽

10.3390/bios11020032 ◽

2021 ◽

Vol 11 (2) ◽

pp. 32

Author(s):

Tamás Gerecsei ◽

Péter Chrenkó ◽

Nicolett Kanyo ◽

Beatrix Péter ◽

Attila Bonyár ◽

...

Keyword(s):

Dissociation Constant ◽

Mammalian Cells ◽

Single Cells ◽

Relevant Information ◽

Mass Action ◽

Optical Data ◽

Live Cells ◽

Label Free ◽

Local Flow ◽

Cell Assays

The binding of integrin proteins to peptide sequences such as arginine-glycine-aspartic acid (RGD) is a crucial step in the adhesion process of mammalian cells. While these bonds can be examined between purified proteins and their ligands, live-cell assays are better suited to gain biologically relevant information. Here we apply a computer-controlled micropipette (CCMP) to measure the dissociation constant (Kd) of integrin-RGD-binding. Surface coatings with varying RGD densities were prepared, and the detachment of single cells from these surfaces was measured by applying a local flow inducing hydrodynamic lifting force on the targeted cells in discrete steps. The average behavior of the populations was then fit according to the chemical law of mass action. To verify the resulting value of Kd2d = (4503 ± 1673) 1/µm2, a resonant waveguide grating based biosensor was used, characterizing and fitting the adhesion kinetics of the cell populations. Both methods yielded a Kd within the same range. Furthermore, an analysis of subpopulations was presented, confirming the ability of CCMP to characterize cell adhesion both on single cell and whole population levels. The introduced methodologies offer convenient and automated routes to quantify the adhesivity of living cells before their further processing.

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Unveil early-stage nanocytotoxicity by a label-free single cell pH nanoprobe

The Analyst ◽

10.1039/d0an01437k ◽

2020 ◽

Vol 145 (22) ◽

pp. 7210-7224

Author(s):

Qingbo Yang ◽

Alexandre Cristea ◽

Charles Roberts ◽

Kun Liu ◽

Yang Song ◽

...

Keyword(s):

Single Cell ◽

Real Time ◽

Early Stage ◽

Single Cells ◽

Label Free ◽

Cell Ph

The developed pH nanoprobe unveiled nanomaterial properties that previously unknown (e.g., devastating cytotoxicity) via real-time label-free monitoring on single cells.

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