Chemistry: molecular biology and microbiology – Apparatus – Bioreactor
Reexamination Certificate
1999-11-30
2001-12-11
Beisner, William H. (Department: 1744)
Chemistry: molecular biology and microbiology
Apparatus
Bioreactor
C435S287100, C435S297500
Reexamination Certificate
active
06329195
ABSTRACT:
BACKGROUND OF THE INVENTION
Cells can be taken from tissues and grown or proliferated extracorporally. Under these conditions, some cell types develop properties which allow unlimited propagation, through permanently repeated mitoses. These cells are denoted as so-called permanent cell lines. Freshly removed cells from tissues (e.g., surgical samples, or tissues from experimental animals) most often do not display these properties. Cells of such so-called primary cultures either lack mitotic activity or perform only a few mitoses, i.e., these cells can only be cultured for a short period of time and only under loss of most of their original functional characteristics (dedifferentiation).
Adhesion of cells to their growth support and the appropriate differentiation of cells is largely dependent on the properties of the growth support (growth substrate).
The introduction of microporous growth supports made up of thin foils of either organic or inorganic materials led to an improvement in differentiation of cells in culture. This is specifically the case for those cells growing as monocellular layers (monolayers) like epithelial and endothelial cells. The culture containers used for that purpose in most cases are hollow cylinders which are sealed on one side by microporous foils on which cells are growing. These cylinders are then placed into larger culture containers in a way that culture medium can reach the cell monolayers from both the apical and the basolateral side (U.S. Pat. No. 4,608,342),the teachings of which are incorporated herein by reference. The culture medium is replaced at certain intervals. Cultures of this type are denoted as “static”. Under these conditions, compounds produced by cells can accumulate in the culture medium and exert negative effects on cell growth and differentiation. For this reason, improved conditions have been developed. Microporous growth substrates can, for example, be placed between two concentric circular mounting unites (cell carrier) and then cells seeded and attached to the growth support. Such cell carriers can then be transferred to a culture medium chamber which is separated by the carrier in two halves. Each half of the chamber can then be perfused with the same medium or media differing in composition. The cell carriers can also be stapled in a specific culture chamber and per(i)fused with culture medium (DE-PS 3923 279), the teachings of which are incorporated herein by reference. These cell culture systems are easy to handle and offer the possibility to grow and/or maintain primary cultures over longer periods of time than under static culture conditions either using solid (plastic, glass) or microporous supports. However, they have the disadvantage that only culture containers for cell carriers having a very small growth area, i.e., about 0.9 cm
2
, are obtainable. The per(i)fusion cell apparatus in which cell carriers can be stacked in order to obtain enough material for biochemical/cell biological analyses has the disadvantage of insufficient oxygenation of the medium between the cell carriers. Furthermore, both devices display mixing inhomogenities during continuous replacement of the growth medium by per(i)fusion.
SUMMARY OF THE INVENTION
The invention relates to a cell culture apparatus or cell culture device, respectively, with at least one growth support for cells, which borders at least on one side to a culture medium space preferably supplied with liquid medium in a continuous mode via inflow and outflow openings.
The present invention relates to a cell culture apparatus which allows the growth and maintenance of cell cultures over prolonged periods of time under conditions which closely resemble the situation within the intact organisms. According to the invention, this can be achieved by implementation of at least one gas-compartment which can be filled with a single gas or a mixture of gases and which is sealed towards the culture medium container by a membrane impermeable to liquids but permeable to gases. This allows that within the culture medium, which is in immediate contact with the cultured cells defined partial pressures (p) of oxygen (O
2
), carbon dioxide (CO
2
), nitrogen (N
2
), or other gases of biological or toxicological significance can be maintained. This is of specific importance, because the cell culture systems utilizing continuous replacement of culture medium available so far do not offer this possibility. None of the systems available ensures the maintenance of constant partial pressures for biologically relevant gases like oxygen or carbon dioxide. Additional advantages and details of the invention are listed below and are further illustrated by the respective figures.
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ACM-Biotech GmbH
Beisner William H.
Hamilton Brook Smith & Reynolds
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