Perfusion bioreactor

In perfusion bioreactors, supernatant is removed from the bioreactor at certain times, the cells are separated from the supernatant, the supernatant is harvested, and the cells are returned to the bioreactor. Perfusion bioreactors can be operated in a variety of modes. The simplest mode is to consistently remove a certain amount of broth each day (i.e., one bioreactor volume/day) and replace with fresh media. This mode is relatively easy to control. However, as the cell density increases, the required nutrient level may not be met. Also,... 

Hosseinkhani, H., Yamamoto, M., Inatsugu, Y, Hiraoka, Y, Inoue, S., Shimokawa, H., and Tabata, Y. 2006b. Enhanced ectopic bone formation using a combination of plasmid Dna impregnation into 3-D scaffold and bioreactor perfusion culture. Biomaterials, 27,1387-1398. 

Cardiac Tissue Requirements Cardiac Tissue Engineering Bioreactor Design Strategies Perfusion Seeding Bioreactors Perfusion... 

Spinner Flask Bioreactor Perfusion Bioreactors for Cylindrical Constructs Perfusion Bioreactors for Anatomically Shaped... 

Spinner Flask, and Rotating Wall Vessel Bioreactors Perfusion Bioreactors Mechanical Loading Bioreactors Surface Shear Bioreactor Limitations and Challenges... 

Chromiak, J. A., J. Shansky, C. Perrone, and H. H. Vandenburgh. 1998. Bioreactor perfusion system for the long-term maintenance of tissue-engineered skeletal muscle organoids. In vitro Cell Dev Biol Anim 34(9) 694-703. 

Recently, there has been success in generating cocultures that more faithfully reproduce in vivo metastatic microenvironments. An ex vivo microscale liver perfusion bioreactor was used to assess metastatic seeding, mimicking the salient features of fluid dynamics and functionality of hepatic parenchyma. Invasion and subsequent growth of breast and prostate carcinoma cells were detected by two-photon microscopy of fluorescently labeled cells. Tumors... 

Naruse et al. proposed another bioreactor design [22,23], in which porcine hepatocyte spheroids are immobilized on non-woven polyester fabric. This device allows more direct contact between hepatocytes and perfused medium and improves, therefore, the mass transfer capacity. The non-woven fabric module expressed better metabohc and synthetic functions at 24 hours than a hollow fiber module containing spheroids in suspension culture. Longer term results are not yet available and the immunoexclusion properties of this fabric have not been addressed. 

Fig. 1. Schematic illustration of a hepatocyte bioreactor with microcarrier-attachted hepatocytes. The capillary membranes are perfused with medium. (Modified from Dixit et al. [29])...

Several innovative membrane-based bioreactor designs have recently been proposed, including that by Sussman et al. [10], which involves the cultivation of hepatoma cells on the exterior surfaces of semipermeable capillary hollow fiber membranes which are bundled together with an enclosing plastic shell (Fig. 2). Required nutrient medium is circulated within the capillaries. After the hepa-tocytes have attached and formed a mass of liver tissue, the capillary membranes are perfused with the media. 

Commercial scale cultivation of mammalian cells is accompHshed using different technologies roller bottles, microcarriers, suspension (batch, fed-batch or perfusion mode) and hollow fiber bioreactors (Table 2). However, especially for products needed in large amounts, suspension cultivation seems to be the most effective system [4, 5]. Suspension-based systems are characterized by a homogeneous concentration of cells, nutrients, metabolites and product, thereby facilitating scale-up [6] and enabling an accurate monitoring and control of the culture [7]. 

Suspension systems can be operated in different modes batch, fed-batch, chemostat, and perfusion (Fig. 1). These operation modes differ basically in the way nutrient supply and metabolite removal are accomplished, which in turn determines cell concentration, product titer and volumetric productivity that can be achieved [8]. The intrinsic limitation of batch processes, where cells are exposed to a constantly changing environment, limits full expression of growth and metabolic potentials. This aspect is partially overcome in fed-batch cultures, where a special feeding strategy prolonges the culture and allows an increase in cell concentration to be achieved. In perfusion and chemostat processes nutrients are continuously fed to the bioreactor, while the same amount of spent medium is withdrawn. However, in perfusion cultures the cells are retained within the bioreactor, as opposed to continuous-flow culture (chemostat), which washes cells out with the withdrawn medium [9]. 

The use of hydrocyclones for separating mammalian cells from the culture medium opens the possibility of using them to perform perfusion in bioreactors. As hydrocyclones have no moving parts, they are ideally suited for operation under aseptic conditions as required by the biotechnology industry. 

Furthermore, for such application, they would require no maintenance, which would avoid additional risks of contamination and allow a continuous operation of the perfusion bioreactor for several months. 

The terminal settling velocity v, can be obtained by Eq. (1) and, in a perfusion system, the overflow rate Q is equal to the product between the specific perfusion rate D and the bioreactor volume V. Hence ... 

Himmelfarb et al. [80] introduced spin-filters as a cell retention device for high cell density perfusion cultivations of mammalian cells in suspension. Spin-filters are cylinders with a porous wall, which are placed inside stirred tank bioreactors, either mounted on the impeller shaft or driven by an independent motor (Fig. 5). Perfusate is pumped out from inside the spin-filter at the same... 

In a spin-filter based stirred bioreactor, there are various forces acting on both the liquid medium and the cell particles gravity force, axial force due to impeller rotation, centrifugal force created by the spin-filter rotation, and radial force (drag) generated by the perfusion flux. 

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