1. FIGURE 26.1 Large Stirrer Flask. Large stirrer flask (5 L) on a magnetic stirrer. Note the offset pendulum that makes an excursion in the annular depression in the base of the flask. The side arms are for sampling or perfusion with CO 2 in air, which would be required with this volume of medium (see Fig. 26.2) (Techne, courtesy of Sterilin.)

2. FIGURE 26.2 Large Stirrer Culture. Diagram of a large stirrer flask suitable for volumes up to 8 L.

3. FIGURE 26.3 Biostat. A modification of the suspension culture vessel in Fig. 26.1, with continuous matched input of fresh medium and output of cell suspension. The objective is to keep the culture conditions constant rather than to produce large numbers of cells (see also Figs. 26.17–26.19). Bulk culture, per se, is best performed in batches in the apparatus in Fig. 26.1 or Fig. 26.17.

4. FIGURE 26.4 Controlled Bioreactors. (a) Stirred bioreactor with paddle, inlet and outlet ports, gas sparging, and control sensors. (b) Air lift bioreactor consisting of two concentric cylinders, with the inner cylinder being shorter at both ends than the outer, thereby creating an outer and an inner chamber. The bottom of the inner chamber carries a sintered steel ring through which 5% CO 2 in air is bubbled. The bubbles rise, carrying the cell suspension with them. Air/CO 2 is vented from the top, and displacement ensures the return of the cell suspension down the outer chamber. (Modified from Griffiths, 2000.)

5. FIGURE 26.5 Large-Scale Bioreactors. (a) Autoclavable reusable RightModel bioreactor, 150-L capacity. (Courtesy of New Brunswick Scientific.) (b) Single use XDR bioreactor where the culture bag is sterile and disposable; capacity 200 to 2000 L. (Courtesy of Xcellerex.)

6. FIGURE 26.6 Wave Bioreactor. A culture bag with a cell suspension in medium is rocked inside a rocking incubator enclosure, maintaining agitation of the cell suspension and enhancing gas exchange. Medium can be fed to the culture bag, regulated by the control unit, and cell product can be harvested to the receiver; both media supply and receiver can be media bags instead of bottles. (Courtesy of Sartorius Stedim.)

7. FIGURE 26.7 BelloCell Aerator Culture. Cells are grown on carriers that are alternately submerged in medium and exposed for gas exchange by the pump action of the lower bellows compartment. (a) Four bioreactors on drive consol with control unit at front. Individual bioreactors at front show the BioNOC carriers cell compartment above central ring and concertina shape of bellows compartment. (b) Connected to reservoirs and peristaltic pump for continuous perfusion. (See also Plate 22d.) (Courtesy of Bellco, Inc.)

8. FIGURE 26.8 Hollow Fiber Perfusion. Sectional diagram of hollow fiber perfusion system. Medium is circulated from a reservoir to the culture chamber by a peristaltic pump. Aeration and CO 2 exchange are via a gas-permeable tubing coil when the assembly is housed in a CO 2 incubator (see also Fig. 7.11.)

9. FIGURE 26.9 Corning Hyperflask. Multisurface flask with 1720-cm 2 growth area and the same footprint as a 175-cm 2 flask. Arrow indicates air space between growth surfaces where gas exchange occurs with growth compartments. (Flask provided by The Automation Partnership.)

10. FIGURE 26.10 Multisurface Propagators. (a, b) Nunc Cell Factory; stacked culture trays, each of 632 cm 2. (a) Two-chamber, 1264 cm 2, and 10-chamber, 6320 cm 2, cell factories. (b) Forty-chamber cell factory, 25,280 cm 2. (Courtesy of Nunc A/S.) (c–e) Corning CellStack. (c) Single-chamber, 636 cm 2, (d) 10-chamber, 6360 cm 2, and (e) forty-chamber, 254,440 cm 2, cell factories. (Courtesy of Corning Life Sciences.)

11. FIGURE 26.11 Filling Nunc Cell Factory.

12. FIGURE 26.12 Corning CellCube. Multisurface cell propagator with 6500-cm 2 available growth surface. (Courtesy of ATCC.)

13. FIGURE 26.13 Roller Culture Bottles on Racks. (a) Small bench-top rack. (Courtesy of Bellco.) (b) Large freestanding extendable rack. (Bellco; Courtesy of Beatson Institute.)

14. FIGURE 26.14 Roller Bottle Culture. The cell monolayer (dotted line) is constantly bathed in liquid but is submerged only for about onefourth of the cycle, enabling frequent replenishment of the medium and rapid gas exchange.

15. FIGURE 26.15 Roller Drum Apparatus. New Brunswick Scientific’s TC-7 Roller Drum. Roller drums are used for roller culture of large numbers of small bottles or tubes. From an original idea by George Gey. (Courtesy of New Brunswick Scientific.)

16. FIGURE 26.16 Cytopore Microcarriers. Porous microcarriers viewed by scanning EM. (a) Bead alone; (b) bead coated with CHO cells. (Courtesy of GE Healthcare.)

17. FIGURE 26.17 Fixed-Bed Reactor. Cells grown on the surface of beads or macrocarriers are perfused with medium. The beads may be glass, plastic, or porous ceramic and are settled in a dense bed resting on a perforated base at the bottom of the culture vessel or, if of a lighter material, are restrained within a cage. Once the culture is established, the beads do not move, and medium percolates around them.

18. FIGURE 26.18 Celligen  310 Bioreactor. (a) Bioreactor chamber with FibraCel  macrocarriers in medium surrounded by water jacket connected to pump and control unit. (b) Impeller, and basket with FibraCel  disks lying alongside. (Courtesy of New Brunswick Scientific.) ® ®

19. FIGURE 26.19 Bioreactor Process Control. Schematic representation of a paddle-stirred bioreactor with direct-reading probes on the left, feeding to a control unit (top left) that stores the data and also regulates conditions within the bioreactor. A sampling port on the right withdraws the cell suspension from the bioreactor for analysis.

20. FIGURE 26.20 Analysis by NMR. Cell growth in hollow fibers, analyzed by NMR. [31P] NMR spectrum of CHO cells growing in a hollow fiber reactor. The cells had been grown on macroporous beads in the extracapillary space of a specially constructed hollow fiber cartridge that could be accommodated within a 25-mm-diameter NMR probe. The cells were present at a density of approximately 7 x 10 7 cells/mL. Abbreviations: PME, phosphomonoesters, including phosphocholine and phosphoethanolamine; P i, extracellular inorganic phosphorus; P i (acid), an acidic (pH 6.7) extracellular P i pool within the bioreactor; PCr, phosphocreatine; γ-ATP, γ-phosphate of ATP; α-ATP, α- phosphate of ATP; PDE, phosphodiesters; DPDE, diphosphodiesters, including UDPglucose; β-ATP, β- phosphate of ATP; NAD(H), nicotinamide-adenine dinucleotide (reduced). The chemical shift scale is referenced to phosphocreatine at 0.0 ppm. (See also Fig. 25.10) (Courtesy of Dr. Kevin Brindle.)

21. FIGURE 26.21 Robotic Cell Culture. The CompacT SelecT cell culture robotic system. (a) Front aspect: (1) incubator chamber with flasks on top left of carousel; (2) main handling chamber with robotic arm in center, below left of number, capping/uncapping and dispensing unit to left of number and pipette container below number; (3) plate incubator chamber; (4) peristaltic pumps for media additions; (5) Cedex cell counter (Innovatis); (6) multiwell plate liquid handling unit. (See also Plate 24.) (b) Flask in gripper of robotic arm, showing flask being rocked, though it can also be shaken from side to side. (c) uncapping, where cap remains on device until flask is returned for recapping. (d) Pipetting cell suspension from front flask to rear flask. (Courtesy of The Automation Partnership.)