Cell migration has a key role in biological processes e.g. malignancy, wound healing, immune response and morphogenesis. Studying migration and factors that influence it is beneficial e.g. for developing drugs to suppress metastasis, heal wounds faster or enhance the response to infection. Though the majority of the literature describes two-dimensional (2D) migration studies in culture dishes, a more realistic approach is to study migration in three-dimensional (3D) constructs. However, simple-to-implement, straight-forward standardized quantitative techniques for analysis of migration rates of cell colonies in 3D are still required in the field. Here, we describe a new model system for quantifying directional migration of colonies in a hyaluronic acid (oxi-HA) and adipic acid dihydrazide (ADH) hydrogel-based 3D matrix. We further demonstrate that our previously-reported image processing technique for measuring migration in 2D (Topman et al., 2011; 2012) is extendable for analyzing the rates of migration of cells that directionally migrate in the hydrogel and are fluorescently-stained with a 4’,6-diamidino-2-phenylindole (DAPI) nuclear stain. Together, the present experimental setup and image processing algorithm provide a standard test bench for measuring migration rates in a fully-automated, robust assay which is useful for high-throughput screening in large-scale drug evaluations, where effects on migration in a 3D matrix are sought.
