3D Cell Culture: A Review of Current Approaches and Techniques

Abstract

Cell culture in two dimensions has been routinely and diligently undertaken in thousands of laboratories worldwide for the past four decades. However, the culture of cells in two dimensions is arguably primitive and does not reproduce the anatomy or physiology of a tissue for informative or useful study. Creating a third dimension for cell culture is clearly more relevant, but requires a multidisciplinary approach and multidisciplinary expertise. When entering the third dimension, investigators need to consider the design of scaffolds for supporting the organisation of cells or the use of bioreactors for controlling nutrient and waste product exchange. As 3D culture systems become more mature and relevant to human and animal physiology, the ability to design and develop co-cultures becomes possible as does the ability to integrate stem cells. The primary objectives for developing 3D cell culture systems vary widely - and range from engineering tissues for clinical delivery through to the development of models for drug screening. The intention of this review is to provide a general overview of the common approaches and techniques for designing 3D culture models.

Quantitative Validation of the Presto Blue™ Metabolic Assay for Online Monitoring of Cell Proliferation in a 3D Perfusion Bioreactor System

“…A further increase in PB solution volume would even allow the further broadening of the window of operations, thereby enabling the methodology to effectively cover the entire range of cell densities used for tissue engineering applications, for which final cell densities in the engineered constructs were reported between 10 7 and 10 8 cells/cm 3 . 9,10,16,31,39 In conclusion, we can state that the PB assay is a promising tool for online monitoring of cell proliferation in a 3D perfusion bioreactor system. Since the conversion of the metabolic assay was shown to be constant in function of time during cell proliferation, and the culture parameters such as flow rate and shear stress did not influence the metabolic activity or the conversion efficiency, a quantitative correlation between the cell number and PB conversion could be established.…”

“…[3][4][5] The integration of bioreactor systems in current laboratory-scale processes therefore holds promise for the translation in a clinical and ultimately commercial setting. 6,7 Bioreactors have been employed frequently to provide sufficient nutrient and oxygen transport and removal of waste products 6,[8][9][10][11][12] while allowing for monitoring and control of physicochemical and biological parameters 7,[13][14][15][16][17] during cell proliferation, differentiation, and the development of cell-carrier constructs. Furthermore, these parameters could be used as nondestructive quality indicators of the cells or the developing construct.…”

Stromal-Cell-Derived Extracellular Matrix Promotes the Proliferation and Retains the Osteogenic Differentiation Capacity of Mesenchymal Stem Cells on Three-Dimensional Scaffolds

“…Researchers utilize a variety of scaffolds to study the behavior of MSCs in 3D cultures. [29][30][31][32][33][34] Typically, these 3D scaffolds are composed of foreign materials (either natural or synthetic) and often elicit an atypical cellular response. Therefore, to study the behavior of MSCs in vitro, an ideal 3D environment should be composed of materials that mimic the native tissue.…”

“…That is, a scaffold should exhibit similar mechanical, architectural, and compositional properties as exhibited in the in vivo bone marrow niche. 26,29 In our previous work, the design and fabrication of a biomimetic collagen/hydroxyapatite (Col/HA) composite scaffold was described. 35 The Col/HA scaffold exhibited properties similar to that of trabecular bone in terms of tissue architecture and composition.…”

Comparison of Microtissue Forming Capacity of Sh-Sy5y and Sk-N-as Cell Lines

“…Scaffold-free cultures are advantegous in providing self-assembly of cells and true physiological interactions between different types of cells without any secondary interfering material [12][13][14]. One of the scaffold-free methods is the micromolded technique utilizing nonadhesive hydrogels, cells of either one type or of more than one type spontaneously aggregate to form 3D microtissue spheroids [12][13][14][15][16][17][18][19] providing physiologically relevant conditions and a suitable platform for testing of drugs at early stage development [16] and high throughput screening processes [9]. In this context, multi-layers of cells within the microtissue spheroid forms natural barriers to drugs as intercellular tight junctions bind cells together and block or slow down the diffusion of drugs as in human tissues [19].…”