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We describe a chip-based platform for the three-dimensional cultivation of cells in micro-bioreactors. One chip can house up to 10 Mio. cells that can be cultivated under precisely defined conditions with regard to fluid flow, oxygen tension etc. in a sterile, closed circulation loop.
We have developed a chip-based cell culture system for the three-dimensional cultivation of cells. The chip is typically manufactured from non-biodegradable polymers, e.g., polycarbonate or polymethyl methacrylate by micro injection molding, micro hot embossing or micro thermoforming. But, it can also be manufactured from bio-degradable polymers. Its overall dimensions are 0.7 1 x 20 x 20 x 0.7 1 mm (h x w x l). The main features of the chips used are either a grid of up to 1156 cubic micro-containers (cf-chip) each the size of 120-300 x 300 x 300 μ (h x w x l) or round recesses with diameters of 300 μ and a depth of 300 μ (r-chip). The scaffold can house 10 Mio. cells in a three-dimensional configuration. For an optimal nutrient and gas supply, the chip is inserted in a bioreactor housing. The bioreactor is part of a closed steril circulation loop that, in the simplest configuration, is additionaly comprised of a roller pump and a medium reservoir with a gas supply. The bioreactor can be run in perfusion, superfusion, or even a mixed operation mode. We have successfully cultivated cell lines as well as primary cells over periods of several weeks. For rat primary liver cells we could show a preservation of organotypic functions for more than 2 weeks. For hepatocellular carcinoma cell lines we could show the induction of liver specific genes not or only slightly expressed in standard monolayer culture. The system might also be useful as a stem cell cultivation system since first differentiation experiments with stem cell lines were promising.
This paper describes the use of a chip-based platform (fig. 1) for the three-dimensional cultivation of cell lines as well as primary cells. Since many cells do express organotypic functions only in a 3D-environment, we have developed a polymer chip that provides a scaffold to which the cells can adhere in all spatial directions, and that can be mounted in a bioreactor housing for the control of fluid flow, oxygen tension etc. Depending on the experimental design, the surface of the polymer can be modified by various tec.......
We have developed a chip-based platform for the three-dimensional cultivation of cells in actively perfused micro bioreactors. The chips can be manufactured from non-biodegradable as well as biodegradable polymers by micro injection molding, hot embossing as well micro thermoforming techniques 3. Depending on the experimental design, the surface of the polymer can be modified by UV-irradiation 4. Hepatocyte cell lines as well as primary rat hepatocytes can successfully be cultivated in these devices.......
We would like to thank Mechthild Herschbach and Anke Dech for excellent technical assistance.
....Material Name | Type | Company | Catalogue Number | Comment |
---|---|---|---|---|
Name | Company | Catalog Number | Comments | |
Cells | Other | ATCC | HB-8065 | |
Collagen I from rat tail | Reagent | Roche Diagnostics | 11 179 179 001 | |
PARIS kit | Reagent | Ambion Inc. | AM1921 | |
Syto16 | Reagent | Invitrogen | S7578 | |
anti cytokeratin 18 | Antibody | Abcam plc | ab668 | Primary Ab, Mouse monoclonal, used 1/100 in PBS |
Anti E-cadherin | Antibody | Abcam plc | ab1416 | Primary Ab, Mouse monoclonal, used 1/50 in PBS. |
Goat anti-albumin | Reagent | Bethyl Laboratories | E80-129 | Primary Ab, goat anti-human Albumin, used 1/200 in PBS |
Rabbit anti-mouse IgG1 | Antibody | Invitrogen | A11059 | Secondary Ab, Alexa Flour 488 conjugated, used 1/100 in PBS + 0.5 % BSA |
Cy3 anti-goat IgG | Reagent | Jackson ImmunoResearch Lab | 705-165-003 | Cy3 AffiniPure donkey a-goat IgG Ab, used 1/700 in PBS + 0.5% BSA |
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