Microfluidics and Single-Cell Technologies

We offer practice-relevant and custom developments of methods and prototypes for processing and handling complex biological specimens. One key focus is the manipulation of individual materials, such as non-invasive and versatile handling of individual cells and, in particular, small cell specimens in microfluidic chips. This primarily involves the use of electrical fields within the radio frequency range, which are absolutely harmless for the cells. This is combined with complementary manipulation processes, such as laser tweezers or microfluidic processes, for more complex tasks. Along with this, we are also concentrating on integrating sensor technology into microfluidic components to obtain data on the key characteristics of cells and other complex biological specimens.

Single-Cell Handling

The Working Group can draw on years of experience in the field of handling valuable cell samples at single-cell level. With the aid of freely configurable microelectrodes that are integrated into microfluidic systems, specific individual cells can be selected from a suspension. The selected cells can be sorted and transferred into other media without centrifugation, and controlled contact between the cells can be triggered for purposes of fusion or signal transfer. This is of interest for immunological cell activation or during the initiation of stem cell differentiation. The quantification of bonding dynamics between two cells or microscale objects is also possible using our technology.

Methods

  • Design and setup of chip-based microsystems
  • Numerical modelling of microsystems using the finite element method
  • Washing, characterizing, sorting and stimulating rare and valuable cell specimens (e.g. stem cells, circulating tumor cells)
  • Non-contact handling of individual cells (e.g. cell selection, cell fusion, cell rotation, cell stimulation, cell characterization, etc.)
  • Cloning of previously characterized or processed single cells
     

Equipment

  • Variable microfluidic setup
  • Computer-controlled pump systems
  • 32-channel radio frequency generators for dielectrophoretic particle manipulation
  • Laser tweezers with a combined UV laser for laser cutting

  • GeSiM Gesellschaft fuer Silizium-Mikrosysteme mbH
  • Surflay Nanotec GmbH
  • NanoBioAnalytics
  • University of Rostock, Chair for Biophysics
  • Tel Aviv University, OMNI Group
  • microfluidic ChipShop GmbH
  • The Fraunhofer Institute for Applied Polymer Research, Life Science and Bioprocesses Research Unit

  • Habaza M, Kirschbaum M, Guernth-Marschner C, Dardikman G, Barnea I, Korenstein R, Duschl C, Shaked NT. Rapid 3D Refractive-Index Imaging of Live Cells in Suspension without Labeling Using Dielectrophoretic Cell Rotation. Adv. Sci. (2017), 4, 1600205
  • Guernth-Marschner C, Kirschbaum M, Jaeger MS, Duschl C. Electrofusion of cells in microdevices. Cell News. (2013), 39(3), 14-18.
  • Kirschbaum M, Gürnth-Marschner CR, Cherré S, de Pablo Peña A, Jäger MS, Kroczek RA, Schnelle T, Müller T, Duschl C. Highly controlled single-cell electrofusion in dielectrophoretic field cages. Lab on a Chip. (2012), 12, S. 443-450.
  • Guido I, Xiong C, Jaeger MS, Duschl C. Microfluidic system for cell mechanics analysis through dielectrophoresis. Microelectron Eng. (2012), 97:379-382
  • Boettcher M, Schmidt S, Latz A, Jaeger MS, Stuke M, Duschl C. Filtration at the microfluidic level: enrichment of nanoparticles by tunable filters. J Phys Condens Mat. (2011), 23, 324101
  • Guido I, Jaeger MS, Duschl C. Dielectrophoretic stretching of cells allows for characterization of their mechanical properties. Eur Biophys J. (2011), 40:281-288.
  • Guido I, Jaeger MS, Duschl C. Influence of medium consumption on cell elasticity. Cytotechnology. (2010), 62, 257-263.
  • Kirschbaum M, Jaeger MS, Duschl C. Correlating short-term Ca2+ responses with long-term protein expression after activation of single T cells. Lab Chip. (2009), 9, 3517-3525.
  • Kirschbaum M, Jaeger MS, Schenkel T, Breinig T, Meyerhans A, Duschl C. T cell activation on a single-cell level in dielectrophoresis-based microfluidic devices. J Chromatogr A. (2008), 1202, 83–89.
  • Böttcher M, Jäger MS, Kirschbaum M, Müller T, Schnelle T, Duschl C. Gravitation-driven stress-reduced cell handling. Anal Bioanal Chem. (2008), 390, 857-863.
  • Storn V, Kirschbaum M, Schlosshauer B, Mack AF, Fricke C. Electrical stimulation-induced release of beta-endorphin from genetically modified neuro-2a cells. Cell Transplant. (2008), 17(5):543-8
  • Jaeger MS, Uhlig K, Schnelle T, Mueller T. Contact-free single-cell cultivation by negative dielectrophoresis. J Phys D Appl Phys. (2008), 41:175502.
  • Jaeger MS, Mueller T, Schnelle T. Thermometry in dielectrophoresis chips for contact-free cell handling. J Phys D Appl Phys. (2007), 40:95–105
  • Böttcher M, Jäger MS, Riegger L, Ducrée J, Zengerle R, Duschl C. Lab-on-chip-based cell separation by combining dielectrophoresis and centrifugation. BRL. (2006), 1(4):443-451.

Non-contact rotation of a mammalian cell in a dielectrophoresis-based microfluidic system. The cell is initially captured in a dielectric field cage by means of dielectrophoretic forces. The cage consists of eight microelectrodes (shown in black): four at the base, and four on the top of the microchannel. By applying radio frequency fields phase-shifted by 90° onto these eight electrodes, the cell can be rotated by any geometrical axis required. Scale bar: 10 µm

Single-cell infusion of a P3X myeloma cell with a B-cell in a dielectric field cage. Individually selected cells are initially captured or paired using dielectrophoretic forces, which allows their cell membranes to be brought into close contact. The cell fusion is then initiated by application of a brief voltage pulse (indicated by a flash of light in the video). Around two minutes after application of the pulse, the image of the cell membranes begins to blur at the point of contact. Around three minutes after application of the pulse, any membrane sections separating the cells have disappeared altogether, allowing one individual heterokaryon to form.

Non-contact handling of individual cells in a dielectrophoresis-based microfluidic system. Deflector elements and dielectrophoretic switches placed in succession (shown in black in the video; active elements are indicated by a white line) are used to divert individually selected cells (arrows) from their original trajectories and transfer them into a zig-zag shaped retaining element perpendicular to the flow lines in the channel. At this point, they are held against the flow of fluid in the channel, making it possible for active substances, or staining and washing solutions, to flow over them.

A contact between one individual T-cell and a bioactive coated microparticle is produced in a dielectrophoresis-based microfluidic system. Individually selected cells or particles are transferred to a zig-zag shaped retaining element using dielectrophoresis, where they are held against the flow of fluid in the channel using a non-contact method. By transferring the cell and particle into the same retaining position, a contact is produced between the objects, which thereby stimulates the T-cell.

Microparticles as Sensors for Bioanalysis

Microparticles as Sensors for Bioanalysis
© Fraunhofer IZI-BB

Accumulation of microparticles in a microchannel by means of dielectrophoresis. The particles are held back against the flow of fluid (red arrow) using a non-contact method, and can be released again at the touch of a button.

Most bioanalytical methods still require excessively complicated and expensive components and equipment. This project is developing dielectric microparticles as sensors for biomolecules on the basis of so-called »whispering gallery modes« (WGM) for use in microfluidic components. This allows the advantages offered by the particles to be used to optimal effect: They are suitable for a wide variety of uses, as they can diffuse freely in the analyte, only require a minimal specimen volume and can also be read using straightforward methods.

Microsystems for Controlling Neuronal Cell Growth

Microsystems for Controlling Neuronal Cell Growth
© Fraunhofer IZI-BB

Neuronal network on a cell culture substrate with a microstructure featuring a thermoresponsive polymer coating. The position of the cell bodies is determined using the geometric structure of the surface coating. The neurites that connect the individual cell groups to each other are clearly visible.

The analysis of artificial neuronal networks is a highly promising approach to addressing a large number of neurobiological areas of inquiry. Despite keen efforts all over the world, a satisfactory solution has yet to be found for in-vitro control over the direction of synaptic transmission between individual cells in a network like this, which, for example, frustrates efforts to explain the form/function relationship in neuronal tissue.

By using microproduction techniques and in close cooperation with our adjoining Working Group, »Microsystems for In-vitro Cell Models«, this project involves the development of cell culture substrates with surface coatings made of thermoresponsive polymers (TRP), which allow neuronal networks with defined connectivity patterns to be produced. The TRP used for this purpose can be transformed from a cell-repellent to a cell-adhesive state according to temperature, which allows the accessibility of a TRP-coated substrate surface for cells and neurite outgrowth to be controlled dynamically and to the exact µm. The objective is to create new methodical approaches to key neuroscientific areas of inquiry in fundamental research, or within the context of the development of pharmaceutical agents.

A Photonic-Microfluidic Production Method for Ultrafast Production of Custom-Made Monoclonal Antibodies

A Photonic-Microfluidic Production Method for Ultrafast Production of Custom-Made Monoclonal Antibodies
© Fraunhofer IZI-BB

Pair of cells in the microfluidic system. The time-lapse sequence shows the highly controlled cell fusion process. The fusion is triggered by applying an electrical pulse to the microelectrode surrounding the pair of cells (shown in black). Scale bar: 10 µm. Time between images: 1 min.

EU Logo

Monoclonal antibodies are one of the binding molecules most frequently used worldwide. A key feature of the standard production method is the (uncontrolled) cell fusion of myeloma and B-cells. This involves the use of complicated selection steps to identify individual antibody-producing cells with the required binding properties among millions of unwanted by-products. This drives the effort and costs of developing antibodies to astronomical heights.

The objective of the project is to establish a method that allows the suitable B-cells to be identified by means of fluorescence optics before fusion occurs, and to use a microfluidic protocol to trigger a controlled fusion with myeloma cells on a single-cell level. This eliminates the need for complex selection steps after the fusion, reducing the work and costs involved to a minimum and allowing tailor-made antibodies to be produced within less than three weeks.

The project is being conducted in close cooperation with the chair for »Physical Chemistry«, and the »Immune Technologies« Working Group at the University of Potsdam, and is receiving financial support from the European Union.