AFM and FluidFM Technologies: Recent Applications in Molecular and Cellular Biology

Abstract

Atomic force microscopy (AFM) is a widely used imaging technique in material sciences. After becoming a standard surface-imaging tool, AFM has been proven to be useful in addressing several biological issues such as the characterization of cell organelles, quantification of DNA-protein interactions, cell adhesion forces, and electromechanical properties of living cells. AFM technique has undergone many successful improvements since its invention, including fluidic force microscopy (FluidFM), which combines conventional AFM with microchanneled cantilevers for local liquid dispensing. This technology permitted to overcome challenges linked to single-cell analyses. Indeed, FluidFM allows isolation and injection of single cells, force-controlled patch clamping of beating cardiac cells, serial weighting of micro-objects, and single-cell extraction for molecular analyses. This work aims to review the recent studies of AFM implementation in molecular and cellular biology.

Figure 1

Atomic force microscopy (AFM). (a) Schematic representation of the AFM. (b) Operation modes of AFM.

Figure 2

SEM and AFM images from skin fibroblasts. (a) SEM image from skin fibroblast (N: nucleus; white bar: 20 μm). (b) Higher magnification of the cytoplasm (white bar: 5 μm). (c) Low-magnification AFM image of fibroblast (➙: cytoskeletal fibers; ∗: AFM artifacts in the form of shadowing structures, +: white bumps; white bar: 10 μm). (d) Higher magnification image showing parallel fibers (white bar: 5 μm). Reprinted with permission from [21], copyright John Wiley and Sons.

Figure 3

Electromechanical signals recorded from a single cardiomyocyte on the MEA chip. (a) The spike train (top) and the synchronized beating pulses (bottom). (b) Zoom on the eletromechanical signals located in the inset frame (black rectangle). Solid signals represent the recording of extracellular field potential, and the dashed ones represent the cardiomyocytes beating force. Reprinted with permission from [3], copyright 2014.

Figure 4

Schematic representation of FluidFM technology. This setup includes a microchanneled cantilever fixed, in a watertight way, to a drilled AFM probe-holder; overpressure can be applied in the nanofluidic channel allowing the delivery of dyes, bioactive molecules, and microorganisms through an aperture in the pyramidal AFM tip. Moreover, the solution contained into the microchannel circuit can also be used to perform electrophysiological experiments. Reprinted with permission from [6], copyright 2009 American Chemical Society.

Figure 5

pc-FluidFM technology. (a) Schematic representation of the pc-FluidFM. (b) An example of pyramidal tip used for the patch clamp experiments. (c) Representative traces of the recorded I_Na current from HEK 293 cells. Adapted with permission from [9], copyright 2015 American Chemical Society.

Figure 6

Schematic representation of the experimental procedure of the single-cell metabolic analysis using FluidFM and MALDI-TOF mass spectrometry. (a) Metabolite sampling using FluidFM. (b) Distribution of the cytoplasmic extract onto a selected MAMS spot. (c) Spraying of the 9AA matrix. (d) Acquisition of MS spectra. Adapted with permission from [31], copyright 2017 American Chemical Society.