Abstract
We describe a novel microfluidic perfusion system for high-resolution microscopes. Its modular design allows pre-coating of the coverslip surface with reagents, biomolecules, or cells. A poly(dimethylsiloxane) (PDMS) layer is cast in a special molding station, using masters made by photolithography and dry etching of silicon or by photoresist patterning on glass or silicon. This channel system can be reused while the coverslip is exchanged between experiments. As normal fluidic connectors are used, the link to external, computer-programmable syringe pumps is standardized and various fluidic channel networks can be used in the same setup. The system can house hydrogel microvalves and microelectrodes close to the imaging area to control the influx of reaction partners. We present a range of applications, including single-molecule analysis by fluorescence correlation spectroscopy (FCS), manipulation of single molecules for nanostructuring by hydrodynamic flow fields or the action of motor proteins, generation of concentration gradients, trapping and stretching of live cells using optical fibers precisely mounted in the PDMS layer, and the integration of microelectrodes for actuation and sensing.
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Abbreviations
- AC/DC:
-
Alternating current/direct current
- AMPPNP:
-
Adenosine 5′-(β,γ-imido)triphosphate
- ASE:
-
Advanced silicon etching
- ATP:
-
Adenosine 5′-triphosphate
- cDNA:
-
Complementary (or copy) DNA
- DNA:
-
Deoxyribonucleic acid
- ECM:
-
Extracellular matrix
- FCS:
-
Fluorescence correlation spectroscopy
- FITC:
-
Fluoresceine isothiocyanate
- FN:
-
Fibronectin
- FRET:
-
Fluorescence resonance energy transfer
- GFP:
-
Green fluorescent protein
- Hz:
-
Hertz (s−1)
- kbp:
-
Kilobase pairs
- μTAS:
-
Micro total analysis system
- nDEP:
-
Negative dielectrophoresis
- nM:
-
Nanomol/liter
- nN:
-
Nanonewton
- OD:
-
Outer diameter
- PCB:
-
Printed circuit board
- PDMS:
-
Poly(dimethylsiloxane)
- PMMA:
-
Poly(methylmethacrylate)
- PNIPAAm:
-
Poly(N-isoproypyl acrylamide)
- POMA:
-
Poly(octadecene-alt-maleic anhydride)
- PTFE:
-
Poly(tetrafluoroethylene)
- RNA:
-
Ribonucleic acid
- TIRF:
-
Total internal reflection fluorescence
- UNF:
-
Universal National Fine Thread
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Acknowledgements
We thank G. Fuhr (Fh-IBMT) and G. Gradl (Evotec Technologies GmbH, Berlin) and their coworkers for cooperation in the field of nDEP and cell sorting, M. Knoblauch (Fh-IME, Aachen) for the supply of forisomes, W. Pompe (TU Dresden) and J. Howard (MPI-CBG Dresden) for useful discussions on nanostructuring, and C. Wenzel and K. Richter (TU Dresden) for their help with ASE. This work was funded by Grants from the German Ministry of Education, Science, Research, and Technology (BMBF) (Grant No. 0314025 and 0314036), the Deutsche Forschungsgemeinschaft (FOR 335), the Saxonian Ministry of Science and Arts (SMWK), and the Sächsische Aufbaubank (SAB) (Grant No. 6988/1099 and 9890/1519).
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Gast, F.U., Dittrich, P.S., Schwille, P. et al. The microscopy cell (MicCell), a versatile modular flowthrough system for cell biology, biomaterial research, and nanotechnology. Microfluid Nanofluid 2, 21–36 (2006). https://doi.org/10.1007/s10404-005-0047-6
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DOI: https://doi.org/10.1007/s10404-005-0047-6