Cell Biology International ISSN 1065-6995 doi: 10.1002/cbin.10116 METHODOLOGY 3D Arrays for high throughput assay of cell migration and electrotaxis Sanjun Zhao 1,2 , Runchi Gao 1,2 , Peter N. Devreotes 3 , Alex Mogilner 4 and Min Zhao 2 * 1 Laboratory of Regenerative Biology, Key Laboratory of Yunnan for Biomass Energy and Biotechnology of Environment, School of Life Sciences, Yunnan Normal University, Kunming 650500, China 2 Institute for Regenerative Cures, University of California, Davis, School of Medicine, Sacramento California 95616, USA 3 Department of Cell Biology and Anatomy, Johns Hopkins University, School of Medicine, Baltimore Maryland 21205, USA 4 Departments of Neurobiology, Physiology and Behavior, University of California at Davis, Davis California 95817, USA Abstract Cell behaviour in 3D environments can be significantly different from those in 2D cultures. With many different 3D matrices being developed and many experimental modalities used to modulate cell behaviour in 3D, it is necessary to develop high throughput techniques to study behaviour in 3D. We report on a 3D array on slide and have adapted this to our electrotaxis chamber, thereby offering a novel approach to quantify cellular responses to electric fields (EFs) in 3D conditions, in different matrices, with different strains of cells, under various field strengths. These developments used Dictyostelium cells to illustrate possible applications and limitations. Keywords: 3D matrices; array; assay; cell locomotion; electrotaxis; morphology Introduction Cell migration is fundamental to many important biological processes including development, regeneration, wound healing and immune responses (Locascio and Nieto, 2001; Luster et al., 2005). Defects in cell behaviour have serious consequences, including delayed wound healing, ineffective immune responses to infection and mental retardation (Wang et al., 2005; Etienne-Manneville, 2008). Cell culture in 2D planar dishes has provided an important enhancement to our understanding of both the phenomena and mechanisms underlying cell behaviour such as spreading, migration and division. Cell migration and other behaviour in 3D, however, can be significantly different from those seen in 2D cultures (Knight et al., 2000). For example, in a 3D matrix, non-malignant mammary cells form polarized, growth-arrested acinus-like colonies, whereas this phenotype is lost when cells are cultured ex vivo on 2D plastic substrata. Significantly, malignant mammary cells form disorganised, proliferative and nonpolar colonies (Lee et al., 2007). In 2D models, no significant difference was observed between the malignant and non-malignant cell lines, while the 3D migration kinetics of the non-invasive cell line was lower than the migration kinetics of the invasive cell line (Hazgui et al., 2005). Overall cellular signalling pathways and cell morphology are dramatically influenced by 3D culture as opposed to traditional 2D monolayers (Weigelt et al., 2010). Many other cell types behave differently in 3D matrices of different materials (Even-Ram and Yamada, 2005; Zaman et al., 2006; Mandal and Kundu, 2009; Klemke et al., 2010; Tayalia et al., 2011). Because 3D systems more closely mimic the in vivo situation, it is critical to verify important results from 2D cultures in 3D systems. Many different types of 3D matrices have been produced that require efficient experimental techniques to determine their effects on cell behaviour. For example, modification of the matrix composition, electrical charge, density, etc. yields hundreds of thousands of different 3D matrix environments for culturing and transplanting cells (Tibbitt and Anseth, 2009; Bott et al., 2010; Tai et al., 2010; Ehrbar et al., 2011; Galie et al., 2011). Different treatments of cells in 3D with various modalities, such as growth factors, toxic agents and different mechanical and physical properties further increase the experi- mental conditions and pose a challenge for efficient deter- mination of cell behaviour in a large number of situations. *Corresponding author: e-mail: minzhao@ucdavis.edu Cell Biol Int 37 (2013) 995–1002 s 2013 International Federation for Cell Biology