Fundamentals and Applications of FluidFM Technology in Single‐Cell Studies

Abstract

various microfluidic devices have been routinely employed. However, in these approaches, there is either a high chance of damaging the cells while detaching the adherent cells from the surfaces, or isolation of individual cells from the adherent cell culture is not at all possible. Table 1 lists the drawbacks and challenges associated with these cell sorting techniques. Recently, atomic force microscope (AFM) has emerged as a significant tool to study microorganisms and mammalian cells in real time. This is due to its compatibility with the physiological conditions of the cells. [3] AFM allows the bioimaging of cells with various degrees of resolution, from whole cells [4] down to individual molecules. [5] Using force-curve based imaging, Alsteens et al. revealed single bacteriophages extruding from the surface of bacteria, [6] whereas our group was able to image nanostructures termed microvilli on the surface of T cells. [7] With the introduction of high-speed AFMs, Yamashita et al. were able to follow the diffusion of single molecules on the surface of living bacterial cells in real time. [8] Valuable biophysical and structural information can be achieved down to a single-cell level by means of high-resolution imaging and force spectroscopy. [9] In force spectroscopy experiments using AFM, a cell is directly attached to the cantilever apex with a biocompatible glue (e.g., polydopamine (PDA)). However, since the cell is irreversibly bound to the AFM probe, each functionalized cantilever can be utilized with only one cell for manipulation. It is, therefore, challenging to perform serial and rapid measurement of a reasonable number of cells; this makes it a time-consuming and elaborate process to address a statistical distribution. For injecting biomolecules such as dyes, enzymes or nanoparticles into single living cells, various approaches such as nano-fountain probes, [10] nanoneedles, [11] and carbon nanotubes [12] have been used. However, these approaches lack precise handling for local dispensing of fluid and are limited to the handling of very small volumes (on the order of attoliters (aLs) up to 50 femtoliters (fLs)). Moreover, using these approaches, a relatively long period of time is required to perform intracellular injection. With the advancement of technologies, it has become relatively easier to address these problems in a more controlled manner. Conventional AFM has been integrated with microfluidics to overcome these limitations. [13] This bioanalytical tool is known as a fluidic force microscope or FluidFM, which was This review describes the potential of FluidFM technology and its implementation in studying the interface between a single cell (prokaryote or eukaryote) and a surface or a surrounding area. A combination of microfluidics with conventional atomic force microscope (AFM) makes this platform efficient to address challenges associated with various biomolecular systems and biophysical activities down to single-cell levels. Upon regulating the pressure through a microchanneled cantilever v...

“Writing biochips”: high-resolution droplet-to-droplet manufacturing of analytical platforms

“…80 They invented the FluidFM, a device based on a microchanneled cantilever chip fixed to a drilled AFM probe holder hollow tip for continuous dispensing of droplets in liquid environments. Importantly, the presence of a pressure controller allows the use of this system not only for delivering biomolecular inks in liquid environments or solid surfaces or for single-cell biophysical investigations, 81,82 but also to perform minimally invasive sampling from intracellular matrices for analyzing the contents. 83 In the context of synthetic biology, this technology has been very recently applied to the scalable fabrication of phospholipid microarrays in air and liquid.…”

FluidFM for single-cell biophysics

“…In this way, FluidFM integrates the advantages of micropipette and AFM, since FluidFM can not only perform three-dimensional manipulations of single cells, but also tune liquid delivery to single cells with high precision (femtoliter) and perform mechanical measurements on single cells with nanometer spatial resolution, providing new possibilities for the studies at single-cell level. Recently, Saha et al [44] have surveyed the applications of FluidFM on force measurements. Here, the advances of FluidFM in single-cell biophysics (including singlecell manipulations, single-cell force spectroscopy (SCFS), and single-cell electrophysiology) are comprehensively summarized and illustrated with examples highlighting the unique capabilities of FluidFM.…”

Producing Plant Virus Patterns with Defined 2D Structure

“…[22,24] Applying an overpressure to the microchanneled cantilever leads to the deposition of the desired material (liquids or nanoparticles), while the feedback of the scanning force microscope regulates the contact between the nanopipette respectively nanosyringe with the substrate. (In addition to the here-utilized mode of spotting, FluidFM can also be used for a variety of applications, such as immobilizing soft colloidal particles [25,26] or cells [27,28] at the tip for single-particle or single-cell force spectroscopy by an underpressure, producing DOI: 10.1002/pssa.202100259…”

“…Deegan and Marangoni effects are shown to play a role if the spotted droplets are not very small. controlled nanopores to suck material out of single cells [27] or to measure the activity of single ion channels, [29,30] and applying specific drugs or interaction partners at specific sites or into selected single cells.) [27,31] We here report on the formation of specific patterns of genetically modified tomato bushy stunt virus (TBSV) particles.…”

“…controlled nanopores to suck material out of single cells [27] or to measure the activity of single ion channels, [29,30] and applying specific drugs or interaction partners at specific sites or into selected single cells.) [27,31] We here report on the formation of specific patterns of genetically modified tomato bushy stunt virus (TBSV) particles. TBSV has a stable icosahedral shape, a diameter of 30 nm, and shows extreme robustness.…”