Maximize In Vitro Culture Possibilities

From Redmond, et al., "Perfused transcapillary smooth muscle and endothelial cell co-culture--a novel in vitro model," In Vitro Cellular & Developmental Biology--Animal, Volume 31:601-609. Copyright 1995 by the Society for In Vitro Biology. Reproduced with permission of the copyright owner. The Cellmax™ culture system uses hollow fiber bioreactor technology in applications as diverse as secreted protein production, lymphocyte expansion, and cellular co-cultivation. The concept of using artificial capillary beds in a bioreactor chamber fed by a medium perfusion circuit for in vitro cell culture was pioneered in 1972 by Knazek et al. (Science 178:65-67, 1972). The original apparatus supported the growth of human choriocarcinoma cells to tissue-like densities and allowed hormones secreted by the cells to be harvested from the circulating medium without disturbing the cell culture. Almost 20 years after Knazek's original work, Cellco, Inc. (a division of Spectrum Corporation, Laguna Hills, Calif.) commercialized this hollow fiber bioreactor technology for research scale as the CELLMAX® system. The CELLMAX® apparatus is now utilized for applications as diverse as secreted-protein production, lymphocyte expansion, endothelial and vascular smooth muscle cell monoculture, and cellular cocultivation. The design of the technology is meant to mimic growth conditions for cells in vivo. For example, the seeding of microvascular endothelial cells within the capillary fibers and of astroglial cells on the outside of the fibers produces a bioartificial blood-brain barrier system. CELLMAX Artificial Capillary Cell Culture Systems The basic perfusion circuit for the CELLMAX® system consists of a positive-pressure displacement pump circulating medium from a reservoir bottle through an oxygen-carbon dioxide exchange unit prior to the flow of medium through the lumen of each individual fiber of the artificial capillary bundle. The final step of the circuit moves the medium back to the reservoir. Capillary cartridges are available not only in different materials (for example, cellulosic, polypropylene, or polyethylene) customized for specific cell types but also with different fiber pore sizes (4 kD, 10 kD, or 30 kD molecular weight cut-off [MWCO]) for the selective retention of macromolecules of interest. For production of antibodies from a hybridoma culture, for example, cells would be grown in the extracapillary space of a cellulosic-based fiber cartridge with a 30 kD MWCO, allowing waste products and cytokines to diffuse freely into the capillary lumen to be recirculated back to the medium reservoir. Antibody molecules can be retrieved by the removal of a portion of the extracapillary medium through sideport tubing on the cartridge itself. For scale-up, Cellco offers the CELLMAX® BigFoot, for larger cartridges, and the CELLMAX® Quad, which operates up to four independent cultures at one time, including four separate flow rates if appropriate. The CELLMAX® High Flow Quad generates flow rates ideal for laboratories culturing endothelial cells under high levels of shear stress (up to 50 dynes/cm2). All of the CELLMAX® configurations can operate within a standard CO2 incubator, using a specially designed low-voltage motor and an ultrathin power cord. The CELLMAX® culture system offers several advantages over traditional protocols, according to John Cadwell, Cellco's director of sales for cell culture products. Cell cultures can be maintained several months, allowing continuous production and harvesting of antibodies, recombinant proteins, or cytokines. For scale-up production of monoclonal antibodies from hybridoma cultures, the need for ascites fluid production is eliminated, while ascites fluid-like concentrations of antibodies of up to 5 mg/ml of medium and total antibody production of 5-50 mg per harvest (approximately every 2 days) are possible with CELLMAX®. Endotoxin contamination of CELLMAX®-produced antibodies is also 1-2 orders of magnitude below the levels inherent in ascites fluid. The CELLMAX® system can be adapted for use with cells traditionally grown in suspension culture or as adherent cells, letting the investigator choose the pore size of the hollow fiber to retain or exclude particular macromolecules. Cadwell described the hollow fiber as "a way of controlling the effects of cytokines and other autocrine effectors on cells," noting as an example that the growth-inhibiting molecule TGF- ß (27 kD MW) can be eliminated from the cultured cells simply by choosing a cartridge with a 30 kD MWCO pore size. Cellco can provide technical support on how particular cell lines can be cultured in the CELLMAX® system and what applications may be suitable. At the National Cancer Institute's Experimental Immunology Branch, David Winkler instituted the use of the CELLMAX® system as a way to avoid issues of foreign protein contamination during large-scale monoclonal antibody production. Hybridoma culture supernatants containing fetal bovine serum and ascites fluid both require purification of immunoglobulin away from other proteins prior to use in immunoassays. Winkler has successfully grown approximately 60 different hybridoma cell lines in the CELLMAX® cartridges under serum-free conditions, allowing the high-concentration antibodies harvested to be directly used in laboratory protocols. The "low tech" design, without excess gadgetry and the relatively low cost of the unit impressed Winkler, who has also had success isolating recombinant protein products from transfected Chinese hamster ovary cells cultured in the units. Drs. Paul Cahill and Eileen Redmond in the Department of Surgery at Georgetown University Medical Center use the CELLMAX® Quad system in their studies of how hemodynamic forces control vascular smooth muscle cell signaling, migration, and proliferation in the absence or presence of endothelial cells. For Cahill, the CELLMAX® system is superior to cone viscometer, parallel plate, or Flexicell methodologies and is the most physiologic commercial system available. Cahill has successfully grown human, rat, bovine, and murine vascular smooth muscle and endothelial cells in the CELLMAX® cartridges. Using the hollow fiber system with vascular endothelial cells (grown within each fiber) and vascular smooth muscle cells (grown on the outside of each fiber) in both monoculture and coculture, Cahill performs RNA, DNA, and protein analyses on material obtained by lysis of cells directly in the cartridge, as well as migration, proliferation, and biochemical assays on intact cells harvested from the cartridges. Future use of the CELLMAX® technology may include even more applications for tissue engineering models of cell culture, according to Cahill. For more information, on the above systems, please contact Cellco at 800-609-3427 or visit the Spectrum Corporation web site at www.spectrumlabs.com. Author Linda Raab, may be reached at raabx004@maroon.tc.umn.edu.

Maximize In Vitro Culture Possibilities

Interested in reading more?

Become a Member of