Fraunhofer Institute for Cell Therapy and Immunology, Branch Bioanalytics and Bioprocesses IZI-BB
Fraunhofer Institute for Cell Therapy and Immunology, Branch Bioanalytics and Bioprocesses IZI-BB

Physiological and Functional Surfaces for Cell Cultivation

Fraunhofer IZI-BB | Surfaces for cell culture
© Fraunhofer IZI-BB
Control of cell adhesion on thermoresponsive surfaces. At 37 °C, the cells are adherent and spread out on the polymer surfaces. By cooling the surface temperature to 25 °C, the cells detach from their substrate and can be harvested by simple rinsing. This process is reversible, i.e. by reheating to 37 °C, the cells can adhere to the substrate again.

We develop coatings of thermoresponsive polymers for cell culture applications with the aim of effectively and gently controlling cell adhesion. At typical cultivation temperatures, cells adhere and proliferate as on a standard cell culture substrate. If the temperature is reduced by a few degrees, the cells can be detached from these coatings by simple rinsing step. The absence of invasive proteases ensures that cell viability and membrane proteins are not compromised during this critical process step. The polymers can be applied homogeneously or in defined patterns to common cell culture substrates at low cost using simple methods such as spin coating, spray coating, spotting or printing. In addition to use as a cell culture substrate, the polymer coating is suitable for cell tests that allow cell migration to be investigated (e.g. wound healing test) or for establishing co-cultures with defined geometric relationships.

Time-lapse movie of the detachment process of L929 mouse fibroblasts during cooling from 37 °C to room temperature. The cells were cultured on a coated cell culture dish. The coating consists of a thermoresponsive polymer applied via a spray method. Duration of the time-lapse recording: 30 min.

Expand all Close all

Structured thermoresponsive surfaces for local control of cell adhesion

© Fraunhofer IZI-BB

Thermoresponsive microgels can be applied locally to different surfaces using pressure or spotting methods. At 37 °C, the cells grow homogeneously on the entire surface. By cooling the surface temperature to 25 °C, cells selectively detach from their substrate.

When the temperature is increased again, the cells repopulate the thermoresponsive areas. This allows cell migration assays to be performed in a defined and reproducible manner, e.g. also in microfluidic channel.

PartiSens: Particle-based substrates for the gentle expansion of high-quality cell samples

This project focuses on the combination of thermoresponsive surfaces and microcarrier cell culture. 1. the cells can be harvested gently and without effort by cooling down. 2. the yield is increased by enlarging the cell culture surface. 3. by using optical sensor particles as cell culture substrate, the growth rate can be determined and thus the process can be controlled.

Investigation of cell-surface interaction

Visualisation of the adhesion area of fibroblasts on a thermoresponsive cell cultivation substrate during cell detachment triggered by cooling the sample to room temperature. Time-lapse image: duration 1 h.

  • Polyelectrolyte layers (layer-by-layer (LbL) deposition) as reservoirs for biomolecules to control adherent cells, layers (self-assembled monolayers (SAM)) of polymers and biomolecules to improve the biocompatibility of synthetic surfaces
  • Techniques for the preparation of homogeneous and structured coatings: Spin coating, dip coating, spraying, spotting, printing (µ-contact printing)
  • Comprehensive range of methods for non-invasive investigation and characterisation of surfaces and coatings: Contact angle measurement, ellipsometry, surface plasmon spectroscopy (SPR), fluorescence microscopic techniques, fluorescence recovery after photobleaching (FRAP)
  • Time-resolved investigation of cell adhesion on functionalised surfaces using total internal reflection microscopy (TIRFM)

Equipment

  • Contact angle measuring device
  • Vapour deposition system for the production of thin metallic layers (Edwards)
  • Multiscope for imaging ellipsometry, surface plasmon resonance / SPR (Optrel)
  • Atomic Force Microscopy for Biological Applications / Bio-AFM (JPK)
  • TIRF microscopy (Olympus)

  • Flechner M, Schaller J, Stahl M, Achberger K, Gerike S, Hannappel Y, Fu J, Jaeger M, Hellweg T, Duschl C, Uhlig K. Adhesion, proliferation and detachment of various cell types on thermoresponsive microgel coatings. Biotechnol Bioeng. (2022), 1– 12.
  • Uhlig K, Wegener T, Hertle Y, Bookhold J, Jaeger M, Hellweg T, Fery A, Duschl C. Thermoresponsive Microgel Coatings as Versatile Functional Compounds for Novel Cell Manipulation Tools. Polymers (2018), 10, 656.
  • Madaboosi N, Uhlig K, Schmidt S, Vikulina AS, Möhwald H, Duschl C, Volodkin D. A “Cell-Friendly” Window for the Interaction of Cells with Hyaluronic Acid/Poly-l-Lysine Multilayers. Macromol. Biosci. (2017), 1700319.
  • Uhlig K, Wegener T, He J, Zeiser M, Bookhold J, Dewald I, Godino N, Jaeger MS, Hellweg T, Fery A, Duschl C. Patterned thermoresponsive microgel coatings for noninvasive processing of adherent cells. Biomacromolecules (2016),  17, S. 1110-1116.
  • Velk N, Uhlig K, Duschl C, Volodkin D. Mobility of Lysozyme in Poly(L-lysine)/Hyaluronic Acid Multilayer Films. Colloids Surfaces B (2016), 47, S. 343-350.
  • Vikulina AS, Anissimov YG, Singh P, Prokopović VZ, Uhlig U, Jaeger MS, von Klitzing R, Duschl C, Volodkin D. Temperature effect on build-up of exponentially growing polyelectrolyte multilayers. Exponential-to-linear transition point. Phys. Chem. Chem. Phys. (2016), 18, S. 7866-7874.
  • Prokopovic VZ, Duschl C, Volodkin D. Hyaluronic acid/poly-L-lysine Multilayers as Reservoirs for Storage and Release of Small Charged Molecules. Macromo. Biosci. (2015), 15, S. 1357-1363.
  • Vikulina AS, Aleed ST, Paulraj T, Vladimirov YA, von Klitzing R, Duschl C, Volodkin D. Temperature-induced molecular transport through polymer multilayers coated with pNIPAM microgels. Phys. Chem. Chem. Phys. (2015), 17, S. 12771-12777.
  • Paulraj T, Feoktistova N, Velk N, Uhlig K, Duschl C, Volodkin D. Microporous polymeric 3D scaffolds templated by the Layer-by-Layer self-assembly. Macromol. Rapid Comm. (2014) 35, S. 1408-1413.
  • Schmidt S, Uhlig K, Duschl C, Volodkin D. Stability and Cell Uptake of Calcium Carbonate Templated Insulin Microparticles. Acta Biomat. (2014), 10, S. 1423-1430.
  • Uhlig K, Boerner HG, Wischerhoff E, Lutz JF, Jaeger MS, Laschewsky A, Duschl C. On the interaction of adherent cells with thermoresponsive polymer coatings. Polymers. (2014), 6, 1164-1177.
  • Madaboosi N, Uhlig K, Jäger MS, Möhwald H, Duschl C, Volodkin D. Microfluidics as A Tool to Understand the Build-Up Mechanism of Exponential-Like Growing Films. Macromol Rapid Comm. (2012), 33(20), 1775-1779.
  • Madaboosi N, Uhlig K, Schmidt S, Jaeger MS, Möhwald H, Duschl C, Volodkin D. Microfluidics meets soft layer-by-layer films: selective cell growth in 3D polymer architectures. Lab Chip. (2012), 12, S. 1434-1436.
  • Schmidt S, Behra M, Uhlig K, Madaboosi N, Hartmann L, Duschl C, Volodkin D. Mesoporous Protein Particles through Colloidal CaCO3 Templates. Adv. Funct. Mat. (2012) 23, S. 116-123.
  • Uhlig K, Boysen B, Lankenau A, Jaeger MS, Wischerhoff E, Lutz JF, Laschewsky A, Duschl C. On the influence of the architecture of poly(ethylene glycol)-based thermoresponsive polymers on cell adhesion. Biomicrofluidics (2012), 6, S. 024129.
  • Uhlig K, Madaboosi N, Schmidt S, Jäger MS, Rose J, Duschl C, Volodkin D. 3D localization and diffusion of proteins in polyelectrolyte multilayers. Soft Matter (2012),  8, S. 11786-11789.
  • Volodkin VS, Schmidt S, Fernandes P, Larionova NI, Sukhorukov GB, Duschl C, Möhwald H, von Klitzing R. One-step formulation of protein microparticles with tailored properties: hard templating at soft conditions. Adv. Funct. Mat. (2012), 22, S. 1914-1922.
  • Schmidt S, Zeiser M, Hellweg T, Duschl C, Fery A, Möhwald H. Adhesion and Mechanical Properties of PNIPAM Microgel Films and their Potential Use as Switchable Cell Culture Substrates. Adv. Funct. Mat. (2010), 20, S. 3235-3244.
  • Uhlig K, Wischerhoff E, Lutz JF, Laschewsky A, Jaeger MS, Lankenau A, Duschl C. Monitoring cell detachment on PEG-based thermoresponsive surfaces using TIRF microscopy. Soft Matter. (2010), 6, 4262-4267.
  • Kessel S, Müller R, Schmidt S, Wischerhoff E, Laschewsky A, Lutz JF, Uhlig K, Lankenau A, Duschl C and Fery A. Thermoresponsive, PEG-based Polymer Layers: Surface Characterization with AFM Force Measurements. Langmuir (2009), 26, S. 3462–3467.
  • Ernst O, Lieske A, Holländer A, Lankenau A, Duschl C. Tailoring of Thermo-Responsive Self-Assembled Monolayers for Cell Type Specific Control of Adhesion. Langmuir (2008), 24, S. 10259.
  • Wischerhoff E, Uhlig K, Lankenau A, Börner HG, Laschewsky A, Duschl C, Lutz JF. Controlled Cell Adhesion on PEG-based Switchable Surfaces. Angew. Chem. (2008), 47, S. 5666-5668.
  • Ernst O, Lieske A, Jaeger M, Lankenau A, Duschl C. Control of cell detachment in a microfluidic device using a thermoresponsive copolymer on a gold substrate. Lab Chip. (2007), 7, 1322–1329.

 

Patents

  • Duschl C, Lankenau A, Lutz J-F, Laschewsky A, Wischerhoff E, Fuhr GR, Bier F. Substrat, Kultivierungseinrichtung und Kultivierungsverfahren für biologische Zellen. DE 10 2010 012 254 A1. 22. Sept. 2011.
  • Duschl C, Hellweg T, Lankenau A, Laschewsky A, Lutz J-F, Schmidt S, Wischerhoff E. Thermoresponsives Substrat mit Mikrogelen, Verfahren zu dessen Herstellung und Kultivierungsverfahren für biologische Zellen. EP 2 550 352 B1. 22. Sept. 2011.

  • Fraunhofer-Institut für Angewandte Polymerforschung, Potsdam
  • Universität Bielefeld, Bielefeld
  • Surflay Nanotec GmbH, Berlin
  • Ibidi GmbH, München
  • GeSiM mbH, Großerkmannsdorf