The established microfluidic tool for nanomanipulation and single-cell biology
FluidFM probe microscopy (FPM) combines the force sensitivity and positional accuracy of an AFM with FluidFM technology by Cytosurge to allow a whole range of exciting applications in single-cell biology and nanoscience.
Nanosurf has the longest experience providing AFM systems with the FluidFM® add-on, as Cytosurge's initial cooperation partner for this innovative technology - the FlexAFM with FluidFM® system was launched in 2013. FluidFM add-ons are available for the FlexAFM and CoreAFM platforms and a unique integrated FPM solution on the FlexAFM.
Highly accurate pressure, force, and position control with optical sample access
Fully integrated system with user-friendly FluidFM® ARYA operator software
FluidFM® microfluidics control system
Compatible with major inverted microscope brands
Different FluidFM® probes: hollow cantilevers designed for specific applications
FluidFM® micropipettes: tipless cantilevers with opening at the cantilever end
FluidFM® nanopipettes: cantilevers with opening at the tip apex
FluidFM® rapid prototyping probes: cantilevers with closed pyramidal tips, ready for FIB milling
Pioneering research within reach
A tool to conduct original research at the frontiers of science
At the Laboratory of Biosensors and Bioelectronics (ETH Zurich) we have been using the Nanosurf’s Flex-FPM system since the very early days of FluidFM®. Together with Cytosurge, SmartTip and Nanosurf we developed this technology establishing several applications from cell adhesion, colloidal spectroscopy, force-controlled SICM and patch clamp, to local chemical cell stimulation. On the other hand, together with the Institute of Microbiology (ETH Zurich, Prof. Vorholt) we pioneered single-cell protocols like injection, extraction and isolation. Our results are well-received by the scientific community because of their originality stimulating ideas for experiments never attempted before. In this groundbreaking phase the Nanosurf team constantly supported us with interest, competence, and flexibility relying on their long-dated AFM experience.
An integrated solution with optical, force, and fluidic control
Optical cell selection by fluorescence properties (left image). The cantilever was gently placed on the cell under force control and trypsin administration was monitored from a co-deposited fluorescence dye (blue). After release, the cell was picked up with the same cantilever and isolated from the rest by placing it in a separate well, sorted by fluorescence (right image; adapted from Lab Chip (2014) 14, 402‑414 with permission from The Royal Society of Chemistry). Data courtesy: O. Guilaume-Gentil, ETH Zürich, Switzerland.
Innovative and intuitive handling
With its touchscreen interface and predefined experimental workflows, e.g for single-cell injection and extraction or for bacterial and cell adhesion force measurements, the intuitive FluidFM ARYA operator software will guide you through each of your experiments step-by-step.
All FluidFM probes come in sterile blister packs and pre-mounted on plastic carrier clips. Each blister pack has a QR code that can be read with the QR code reader supplied with the Flex-FPM system to allow direct import of detailed cantilever information into the operator software.
Applications
FluidFM application examples
Cell-cell adhesion forces
Recently, FluidFM™ cell adhesion measurements were extended to study cell-cell interaction. This can be the force between a cell (on the cantilever) and a cell below on a substrate (fig. 1 A), but also between a cell and its surrounding cells in a confluent layer (fig. 1 B).
Fig. 1: Cell-cell interactions (red springs) studied by aspiration of single cells to a hollow FluidFM™ probe (orange). A) Probing the force between a cell immobilized on the cantilever and a cell on the substrate, B) picking a single cell from a confluent layer, probing cell-substrate (purple) and cell-cell (red) interactions.
Dr. Noa Cohen of Prof. Tanya Konry's group at Northeastern university in Boston studied cell-cell adhesion forces with a Flex-FPM system to gain more insight into tumor progression and metastasis [Cohen et al. (2017)]. Figure 2 shows an optical image of the method depicted in fig. 2 A that was used by Cohen for this study.
Fig. 2: Optical images showing A) a single cell to be picked up by a FluidFM probe B) the cell aspired to the cantilever and C) the FluidFM probe with aspired cell during a cell-cell adhesion measurement. Data courtesy of Tanya Konry group, Northeastern University, Boston, USA.
Single MCF7 breast cancer cell were immobilized on the cantilever. The cell was then pushed on different cell types that were immobilized on a substrate. The measured cell adhesion between MCF7 cancer cells to different cell types were found to develop differently with incubation time. In these experiments, the reversible binding of cells allowed the different cell pairs to be studied with the same probe (fig. 3), allowîng better comparison of measurements.
Fig. 3 A) Typical force curves between a MCF7 cell aspired to the cantilever and a non-cancerous, fibroblast (HS5) on the substrate at different contact times. B) Development of the force with contact time between the cells. Data courtesy of Tanya Konry group, Northeastern University, Boston, USA.
Dr. Ana Sancho from the group Prof. Jürgen Groll's group at the University of Würzburg extensively studied the interaction between a cell and its neighbors in a confluent layer of epithelial cells (fig. 2 B) [Sancho et al. (2017)]. Fig. 4 shows the cantilever picking up a cell from a confluent layer (A). After removal the empty space from where the cell was picked up is visible (B). Again, the epthelial cells of interest could be selected based on cell size and cell shape using the inverted microscope.
Fig. 4: Confluent layer of cells, where one is pulled out by FluidFM, adapted from: Sancho et al. (2017), Scientific Reports volume 7, 46152.
Human endothelial cells from the umbilical artery were found to exert strong intercellular forces (figures 6 A & B). The forces could be decreased significantly by overexpression of the so-called Muscle Segment Homeobox 1 (MSX1). The MSX1 induces a endothelial-to-mesenchymal transition. This transition is a process involved in cardiovascular development and disease. Complementary to these adhesion experiments, the Flex-FPM system was also used to perform nano-indentation measurements. To this end colloidal beads aspired to the cantilever and force curves were recorded on the cells.
Fig. 5: A) Typical single cell force curves of individual cells or cells in a confluent layer, depicting the increase in adhesion force by cell-cell interactions. B) Effect of MSX1 on the observed cell adhesion for individual cells and cells in a monolayer. Grey and black bars: control measurements on individual cells and monolayers, resp., pale and light blue MSX1 treated individual cells and cells in a monolayer, resp. Adapted from: Sancho et al. (2017), Scientific Reports volume 7, 46152.
Both examples strongly benefitted from FluidFM™ technology provided by the Flex-FPM solution. In case of the confluent layer the large forces of up to over 1.5µN eliminate chemical binding to study cell-cell adhesion forces. In both cases the reversible binding provided the experiments with the necessary speed-up to obtain sufficient statistics.
Spotting and lithography with FluidFM
In cell biology, micropatterning is becoming a widely accepted technique to address cellular behavior at the single-cell level. The ability to guide cell growth by local suppression or stimulation, for example, is beneficial for studying cell growth, for developing cell-based sensors, and for tissue engineering applications. The production of microfabricated biosensors is another application that requires precise placement of biomaterials on a substrate.
FluidFM® technology by Cytosurge enables deposition of (bio)molecules and particles at defined locations with micrometer accuracy and with femtoliter volumes. The closed channel is capable of depositing molecules from a liquid in both air and liquid environment. This enables numerous applications in biomedicine, cell- and microbiology, as well as non-biological nanolithography.
Automatically varying back pressure and contact time in a grid-like pattern (3 dots per condition), spot sizes can be quickly optimized.
Nanosurf logo written in in air with a solution containing approximately 50% glycerol; back pressure 200 mbar.
FluidFM image gallery
Colloidal spectroscopy with FluidFM®
Single Cell Force Spectroscopy
Spotting and lithography with FluidFM
References
FluidFM references
Hear directly from FluidFM® pionieers
For insight on the FluidFM® technology and applications watch this interview with Prof. Dr. Janos Vörös and Dr. Tomaso Zambelli from the ETH Zurich (Switzerland), who were among the first scientists to use fluidic probe microscopy in their research.
Tunable Single-Cell Extraction for Molecular Analyses
O Guillaume-Gentil, RV Grindberg, R Kooger, L Dorwling-Carter, V Martinez, D Ossola, M Pilhofer, T Zambelli, and JA Vorholt
Researchers from ETH Zurich have published extraction of sub-picoliter samples of nucleoplasm and cytoplasm from live cells in the renowned journal Cell. Measurements were performed with the Nanosurf Flex-FPM system that allowed single-cell extraction without killing the cells. The paper describes the method of single-cell extraction as well as the molecular analysis of the extracted samples by TEM, protein assays and PCR. The results illustrate a new FluidFM® application to study cell processes at the single-cell level thus enabling studying the heterogeneity of cells.
Local Chemical Stimulation of Neurons with the Fluidic Force Microscope (FluidFM)
Mathias J. Aebersold, Harald Dermutz, László Demkó, José F. Saenz Cogollo, Shiang-Chi Lin, Conrad Burchert, Moritz Schneider, Doris Ling, Csaba Forró, Hana Han, Tomaso Zambelli and János Vörös
This new stimulation approach, which combines FluidFM for gentle and precise positioning with a microelectrode array read-out, makes it possible to modulate the activity of individual neurons chemically and simultaneously record their induced activity across the entire neuronal network. The presented platform not only offers a more physiological alternative compared with electrical stimulation, but also provides the possibility to study the effects of the local application of neuromodulators and other drugs.
FluidFM: Precise fluidic positioning and delivery platform with applications in cell biology and soft matter
Patrick Frederix, Paul Werten, Dalia Yablon
The platform of fluid force microscopy (FluidFM) offers researchers unique capabilities in precise and well-controlled fluid delivery in small quantities down to femtoliters to the surface. It is based on a microfluidic controller coupled with an innovative AFM probe where a microchannel is hollowed out so that the probe essentially functions as a nanopipette. Applications of FluidFM are described including enhanced manipulation of cells, improved force measurements on biological materials including bacteria, and fluidic writing on hydrogels.
Colloidal Properties of Recombinant Spider Silk Protein Particles
N Helfricht, E Doblhofer, JFL Duval, T Scheibel, and G Papastavrou
Researchers from the group of Prof. Georg Papastavrou at the University of Bayreuth used the Nanosurf Flex-FPM system to perform direct force measurements in the sphere/sphere geometry. Colloidal particles have been prepared from polyanionic and polycationic recombinant spider silk proteins. The amino acid sequences of these spider silk proteins were identical except for 16 residues bearing either cationic or anionic groups, leading to opposite surface charges. Electrokinetic measurements and modelling predicted a soft and porous structure of these protein particles. The presence of a fuzzy, ion-permeable interface has been confirmed by direct force measurements with colloidal particles aspirated reversibly to a FluidFM® probe.
