What is the difference between 2D and 3D Cell Culture?25/03/19
While 2D culture has traditionally been a very stable and successful method of growing simple cells, 3D cell culture is now bringing new capabilities and performance to analysts in the lab. Are you taking advantage of this new approach to biological research? Could 3D culture give your lab new options for analysis, or to improve production?
In this post we take a look at what 2D and 3D cell culture are, the similarities and differences between them and how they can be used in research and production.
2D cell culture – proven, simple but flat
2D cell culture is the conventional approach that most analysts are familiar with and has been in use since the early 1900s. It involves securing, nourishing and growing cell cultures on a flat surface, such as the bottom of a petri dish or flask. For more information please read our introduction to cell culture here.
2D cell culture systems are well established and proven – they form the basis for almost all current routine assays and have a wealth of reference literature. Most existing workflows are built around the 2D culture model, therefore 2D cultures and the associated equipment are relatively inexpensive and easy to use. However, the extensive work with 2D culture systems in both research and production, has shown that there can be issues with the technique as cells in the body do not naturally grow in a 2D fashion.
In the body cells do not grow in a 2D environment, so conditions in a 2D system cannot accurately simulate the conditions the cells would encounter in vivo. Therefore, cells grown in 2D do not give a normal cell morphology.
Gene and Protein Expression
Cells grown in 2D systems often do not show gene and protein expression that is representative of in vivo patterns.
Exposure to Culture
When cells are grown on a flask or petri-dish, all the cells are equally exposed to the culture medium, including any growth factors, additives or drugs that are contained within it. This is not the case for cells in the body – drugs and nutrients are not uniformly spread to all cells. A 2D culture system cannot replicate the diffusion constraints usually found in the body; cells on the outside can readily exchange nutrients and waste products, whereas cells in the centre of the 3D culture cannot make this exchange so freely.
In a 2D system cells can be oversensitive to drugs and treatments, either due to the lack of a diffusion gradient, or because different genes are not expressed as they would be in vivo. The 2D model does not offer an accurate drug response prediction.
2D cell cultures are also relatively inflexible and only a limited range of cellular behaviours can be tested.
These are just some of the reasons why a 3D environment for culture has been gaining traction in recent years.
3D cell culture – a new dimension
3D cell culture has technically been around for a long time. A basic method, known as hanging drop, was tested by Ross Granville (1870 – 1959) and led to advances in a number of areas of biology including oncology and genetics. However it is only in recent years that the technique has become a lot more popular thanks to new cell lines, media becoming available and increased computing, imaging and simulation capabilities being more accessible to analysts.
3D culture conditions more closely resemble the natural environment for cells and so can provide more physiologically useful information that improves cell culture accuracy and flexibility. In order to facilitate cell development in three dimensions a vessel, culture media and, in many cases, a scaffold are required, along with the relevant cell nutrition and incubation conditions. With a vessel and culture conditions capable of supporting cell development and growth in three dimensions, a whole new set of applications can be opened up.
3D cell cultures enable ‘flows’ to be modelled and investigated. In the body cells come into contact with and are affected by the flow of various liquids such as blood or urine. Only in 3D is it possible to replicate these conditions in order to generate target cells or test a growth environment that more closely matches the body. 3D culture also enables barrier tissues to be simulated or assessed and reduces the reliance on animal models by providing more accurate simulations of the natural environment in which cells are found. This increases the accuracy, safety and sustainability of analysis as it is a more direct method of growing and testing cells.
The importance of the 3D scaffold
One of the major differences between the two approaches is the use of a suitable ‘scaffold’ in 3D cell culture. This isn’t used in every process but is increasingly the method of choice for analysts due to its success and versatility. In order to enable cells to grow and proliferate in three dimensions a means of physically supporting them and providing nutrition is needed that works above the base surface of the vessel.
The scaffold more evenly distributes nutrients without restricting passage of cells in applications where cell motion is important. 3D scaffolds can be biomimetic, acting as an extracellular matrix (ECM) formed of materials such as hydrogel or collagen, or can they be bioinert and primarily structural (such as polystyrene). Some 3D applications also use microfluids with moulded microchannels that incorporate other, biomimetic scaffolds. Without the scaffold the entire cell culture would be structurally compromised – it would be difficult to separate target cells for testing or sub-culture, and any infection, corrupted cells or invasive material would spread more quickly through the culture. The scaffold also makes it possible to design more specific physiological environments and improve batch-to-batch consistency of media.
3D cell culture is here to stay and opening up new possibilities for analysts in a wide range of areas when the right culture conditions and scaffold are used. In our recent our recent white paper we explain how our jellyfish collagen can act as an excellent 3D scaffold.
- The Future of Cell Culture, Jellagen
- 3D CELL CULTURE – easier said than done!