Three-Dimensional (3D) in vitro cell culture protocols to enhance glioblastoma research

Abstract

Three-dimensional (3D) cell culture models can help bridge the gap betweenin vitrocell cultures andin vivoresponses by more accurately simulating the naturalin vivoenvironment, shape, tissue stiffness, stressors, gradients and cellular response while avoiding the costs and ethical concerns associated with animal models. The inclusion of the third dimension in 3D cell culture influences the spatial organization of cell surface receptors that interact with other cells and imposes physical restrictions on cells in compared to Two-dimensional (2D) cell cultures. Spheroids’ distinctive cyto-architecture mimicsin vivocellular structure, gene expression, metabolism, proliferation, oxygenation, nutrition absorption, waste excretion, and drug uptake while preserving cell–extracellular matrix (ECM) connections and communication, hence influencing molecular processes and cellular phenotypes. This protocol describes thein vitrogeneration of tumourspheroids using the low attachment plate, hanging drop plate, and cellusponge natural scaffold based methods. The expected results from these protocols confirmed the ability of all these methods to create uniform tumourspheres.