Beyond the paradigm of nanomechanical measurements on cells using AFM: an automated methodology to rapidly analyse thousands of cells

Proa-Coronado, Sergio; Severac, Childérick; Martinez-Rivas, Adrian; Dague, Étienne

doi:10.1039/c9nh00438f

Cited by 13 publications

(32 citation statements)

References 48 publications

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“…A more suitable and scalable approach is based on nanoindentation experiments, exploiting atomic force microscopy (AFM) [29] or recently introduced cantilever-based devices, such as ferrule-top [30], that offer improved usability [31]. The potential of nanoindentation to measure the overall mechanical properties of single cells is nowadays established [32], and the technology is rapidly growing to provide higher experimental throughput [33]. Nevertheless, only few extensions of the analysis have been proposed to explicitly account for the contribution of the AC to cell mechanics [24,34] and none of them emerged as a consolidated and broadly adopted approach.…”

Section: Introductionmentioning

confidence: 99%

Elasticity spectra as a tool to investigate actin cortex mechanics

et al. 2020

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Background The mechanical properties of single living cells have proven to be a powerful marker of the cell physiological state. The use of nanoindentation-based single cell force spectroscopy provided a wealth of information on the elasticity of cells, which is still largely to be exploited. The simplest model to describe cell mechanics is to treat them as a homogeneous elastic material and describe it in terms of the Young’s modulus. Beside its simplicity, this approach proved to be extremely informative, allowing to assess the potential of this physical indicator towards high throughput phenotyping in diagnostic and prognostic applications. Results Here we propose an extension of this analysis to explicitly account for the properties of the actin cortex. We present a method, the Elasticity Spectra, to calculate the apparent stiffness of the cell as a function of the indentation depth and we suggest a simple phenomenological approach to measure the thickness and stiffness of the actin cortex, in addition to the standard Young’s modulus. Conclusions The Elasticity Spectra approach is tested and validated on a set of cells treated with cytoskeleton-affecting drugs, showing the potential to extend the current representation of cell mechanics, without introducing a detailed and complex description of the intracellular structure.

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Section: Introductionmentioning

confidence: 99%

Elasticity spectra as a tool to investigate actin cortex mechanics

et al. 2020

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“…Like optical and spectroscopic features, the mechanical parameters can be considered additional and might be exploited in diagnostic assays and ARTs. Furthermore, due to the enormous potential of such microscopies, numerous efforts are devoted to developing automated systems, such as AFM, which can perform the acquisition of numerous force-distance curves to measure the biomechanical properties of large cell populations or tissues [ 104 ].…”

Section: The Impact Of Afm and Snom Findings On Reproductive Medicinementioning

confidence: 99%

Scanning Probe Microscopies: Imaging and Biomechanics in Reproductive Medicine Research

Andolfi

Battistella

Zanetti

et al. 2021

IJMS

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Basic and translational research in reproductive medicine can provide new insights with the application of scanning probe microscopies, such as atomic force microscopy (AFM) and scanning near-field optical microscopy (SNOM). These microscopies, which provide images with spatial resolution well beyond the optical resolution limit, enable users to achieve detailed descriptions of cell topography, inner cellular structure organization, and arrangements of single or cluster membrane proteins. A peculiar characteristic of AFM operating in force spectroscopy mode is its inherent ability to measure the interaction forces between single proteins or cells, and to quantify the mechanical properties (i.e., elasticity, viscoelasticity, and viscosity) of cells and tissues. The knowledge of the cell ultrastructure, the macromolecule organization, the protein dynamics, the investigation of biological interaction forces, and the quantification of biomechanical features can be essential clues for identifying the molecular mechanisms that govern responses in living cells. This review highlights the main findings achieved by the use of AFM and SNOM in assisted reproductive research, such as the description of gamete morphology; the quantification of mechanical properties of gametes; the role of forces in embryo development; the significance of investigating single-molecule interaction forces; the characterization of disorders of the reproductive system; and the visualization of molecular organization. New perspectives of analysis opened up by applying these techniques and the translational impacts on reproductive medicine are discussed.

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“…Several passes may be necessary to reach a high filling rate. NOTE: A full description of this method is available in refernce 13 . 3.1.5.…”

Section: Sample Preparationmentioning

confidence: 99%

“…Indeed, while the typical method allows one to probe individual cell surface heterogeneity, using our approach, we are able to access the entire cell population heterogeneity. To achieve this objective, we have combined a method consisting in directly immobilizing microbes (here the yeast species Candida albicans) into the wells of a PDMS microstructured stamp 12 , and developing an original program for moving the AFM tip, automatically, from cell to cell 13 and measuring the mechanical properties of each cell. PROTOCOL 1.…”

Section: Introductionmentioning

confidence: 99%

Automation of Bio-Atomic Force Microscope Measurements on Hundreds of <em>C. albicans</em> Cells

Severac

Proa-Coronado

Formosa-Dague

et al. 2021

JoVE

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The method presented in this paper aim at automatizing Bio-AFM experiments and especially the recording of force curves. Using this method, it is possible to record, in 4 hours, forces curves, on 1000 cells, automatically. To maintain a 4 hours analysis time, the number of force curves per cell is reduced down to 9 or 16. The method combines a Jython based program and a strategy for assembling cells on defined patterns. The program, implemented on a commercial Bio-AFM, is able to center the tip on the first cell of the array and then to move, automatically, from cell to cell while recording force curves on each cell. Using this methodology, it is possible to access the biophysical parameters of the cells such as their rigidity, their adhesive properties etc. Thanks to the automation and the large number of cells analyzed, one can access the

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Beyond the paradigm of nanomechanical measurements on cells using AFM: an automated methodology to rapidly analyse thousands of cells

Abstract: This paper reports a methodology which includes an algorithm able to move an AFM tip onto a single cell and through several cells combined with a smart strategy of cell immobilization.

Cited by 13 publications

References 48 publications

Elasticity spectra as a tool to investigate actin cortex mechanics

Elasticity spectra as a tool to investigate actin cortex mechanics

Scanning Probe Microscopies: Imaging and Biomechanics in Reproductive Medicine Research

Automation of Bio-Atomic Force Microscope Measurements on Hundreds of <em>C. albicans</em> Cells

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