Method and devices for the in vitro production of arrangements of cell layers
Inventors
Mosig, Alexander • Rennert, Knut
Assignees
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Publication Number
US-10731119-B2
Publication Date
2020-08-04
Expiration Date
Abstract
The invention is directed to a method for the in vitro production of arrangements of cell layers in which a first well (2.1) which is closed off from its environment except for a first inlet opening (4.1) and a first outlet opening (5.1) and which has as a first cell substrate (6.1) a first wall (2.1.1) and as a second cell substrate (6.2) an opposite, second wall (2.1.2) which is separated from the first wall (2.1.1) by a first gap, a free surface of a cell substrate (6.1, 6.2) to be colonized with cells is oriented orthogonal to the Earth's gravitational force, and cells (9) are adhered to the cell substrate (6.1, 6.2) to be colonized. The invention is further directed to a method of maintaining the biological functionalities of the cell layers and semi-finished products of a device for the in vitro production and culturing of cell layers and a method for the production of the device.
Core Innovation
A method and device are described for in vitro production and culturing of cell layers using semi-finished products in which one or at least two semi-finished products are mounted one above the other. Each semi-finished product has a first and a second well arranged one above the other in a single base plate having an upper side and a lower side, with the first well formed in the upper side and the second well formed in the lower side.
The first and second wells are separated by a single porous membrane with two lateral surfaces, and the semi-finished products are sealed against the environment by a bonding foil on the upper side and a bonding foil on the lower side. A first wall of the first well is formed as a first cell substrate by the bonding foil on the upper side, and a first wall of the second well is formed as a fourth cell substrate by the bonding foil on the lower side.
A second wall of the first well is formed as a second cell substrate through one of the lateral surfaces of the membrane, and a second wall of the second well is formed as a third cell substrate through the other of the lateral surfaces of the membrane. Cell sedimentation by gravity and cell adhesion are achieved by orienting cell substrate surfaces orthogonally to gravity and reorienting the well.
The wells are shaped and connected to inlet and outlet openings to enable controlled fluid flows for perfusion and maintenance of functionality of the cell layers, including first fluid flow in the first well and second fluid flow in the second well through channel-like restrictions. Modular biochip configurations are described that support organoid modeling and measurement of functionality using fluorescence-based contactless measurement points and impedance/electrode setups for evaluating colonization and function or vitality.
Claims Coverage
The independent claim coverage identifies 5 inventive features. It centers on stacked semi-finished products for in vitro production and culturing of cell layers, vertically separated wells divided by a porous membrane and sealed by bonding foils, defined cell substrate walls formed by the bonding foils and membrane lateral surfaces, and inlet/outlet fluid flow arrangements for two different fluid flows.
Stacked semi-finished products for in vitro production of cell layers
One or at least two semi-finished products are mounted one above the other, with a first and a second well arranged one above the other in a single base plate with an upper side and a lower side.
Vertically separated wells divided by a single porous membrane sealed by bonding foils
The first and second wells are separated by a single porous membrane with two lateral surfaces and sealed against the environment by a bonding foil on the upper side and a bonding foil on the lower side.
Cell substrate walls defined by bonding foils and membrane lateral surfaces
A first wall of the first well is formed as a first cell substrate by the bonding foil on the upper side, a first wall of the second well is formed as a fourth cell substrate by the bonding foil on the lower side, a second wall of the first well is formed as a second cell substrate through one lateral surface of the membrane, and a second wall of the second well is formed as a third cell substrate through the other lateral surface of the membrane.
Tapered/rectangular well geometry with inlet and outlet openings for a first fluid flow
The first well narrows to a point at its ends and is rectangular in its middle part, with inlet and outlet openings for ingress and egress of a first fluid flow.
Rectangular second well with inlet/outlet openings and channel-like flow restrictions for a second fluid flow
The second well is rectangular and connected to a second inlet opening and a second outlet opening for ingress and egress of a second fluid flow through channel-like restrictions.
The claim coverage centers on stacked semi-finished products with vertically separated wells divided by a single porous membrane and sealed by bonding foils, with defined cell substrate walls and distinct inlet/outlet opening arrangements for controlled fluid flows in the first and second wells.
Stated Advantages
Cell sedimentation by gravity and cell adhesion are achieved by orienting cell substrate surfaces orthogonally to gravity and reorienting the well.
Controlled fluid flows for perfusion and maintenance of functionality of the cell layers are enabled.
Optical-friendly porous transparent membranes support microscopy.
Fluorescence-based contactless measurement points and impedance/electrode setups are used for evaluating colonization and function or vitality.
Documented Applications
In vitro production and culturing of cell layers.
Organoid modeling.
Measurement of functionality.
Evaluating colonization and function or vitality.
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