Culture apparatus for three dimensional cell growth
Cells that make up tissues in the body possess a complex three-dimensional (3D) architecture which is markedly different from the flat monolayer structure of their cultured counterparts commonly grown on the two-dimensional (2D) surface of tissue culture plastic. The 3D structure of cells and interactions with their neighbours significantly influences their ability to grow and function. Evidence for the functional superiority of cells cultured on 3D supports compared to 2D monolayer cultures has created much interest in the fabrication of materials that promote the growth and function of cells in a more realistic manner.
We have produced a cell culture device that provides an environment for routine 3D cell growth. We have developed thin layers of micro-cellular polymers with a well defined and uniform architecture. Our matrices are made from inert polystyrene using emulsion templating (Barbetta et al. 2005) and their surface chemistry can be varied to influence cell adhesion, proliferation and function (Hayman et al. 2005). By changing the chemical composition of the emulsion and the processing conditions, the porosity of the material can be adjusted to suit alternative applications (Carnachan et al. 2006).
Traditionally, cultured cells normally grow on treated-polystyrene 2D surfaces as in standard cell culture plastic-ware. Using our technology, we can present to cells the equivalent growth substrate but in a 3D format. These materials are readily adaptable to different types of existing tissue culture plastic-ware (e.g. multi-well plates, well inserts). The culture device is pre-fabricated, sterile, is ready to use off-the-shelf and can be handled in a similar manner as standard 2D plastic-ware. These are distinct advantages over some existing 3D supports employed in vitro which are technically more difficult to use, have a finite shelf life and are expensive.
We have exemplified the application of this technology by growing a broad selection of different cell types on these 3D supports and demonstrate enhanced functional activity compared to cells grown under identical conditions on 2D culture plastic. We have also shown that the differentiation of cultured stem cells is significantly influenced by 3D growth (Hayman et al. 2004).
We propose that this technology enables the routine culture of cells in a 3D format in a simple and straightforward way. In preparation to making this patented technology available for use by biotechnologists worldwide, we wish to develop alliances with third parties to further develop this device and plan for its manufacture and production.
Please see below for examples of the application this technology and associated publications. (View relevant publications)
Example application for routine use in cell culture:
The following examples depict the application of styrene-based scaffolds as thin layers adapted for use in existing cell culture vessels such as a multi-welled plate or well insert.
Notes:
- Cell type 1 (red dots) grown as a 3D culture; cell type 2 (green dots) grown as an adherent monolayer
- Dashed line illustrates level of culture medium; hashed pattern illustrates 3D polystyrene scaffold
- Examples 1 & 2 can be combined so that 3D cell growth by two populations of cells occurs in co-culture in the same well
- This concept has been exemplified with >10 different cell types to demonstrate its use as a conventional research tool
Photograph below of prototype well inserts carrying the 90% phase volume polystyrene scaffold at 100mm thick. These examples are of inserts designed to fit into 6-welled (large insert) and 12-welled (small insert) culture plates.
Scanning electron micrograph showing MG63 osteoblasts cultured on 90% phase volume polystyrene scaffolds for 14 days in vitro. The polymer represents the white areas whilst cells (in grey) grow in between.
High magnification scanning electron micrograph showing HEPG2 hepatocytes attached to a strut of three-dimensional styrene polymer for 14 days in vitro.