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An extracellular matrix microarray for probing cellular differentiation

Nature Methods volume 2, pages 119–125 (2005)Cite this article

Abstract

We present an extracellular matrix (ECM) microarray platform for the culture of patterned cells atop combinatorial matrix mixtures. This platform enables the study of differentiation in response to a multitude of microenvironments in parallel. The fabrication process required only access to a standard robotic DNA spotter, off-the-shelf materials and 1,000 times less protein than conventional means of investigating cell-ECM interactions. To demonstrate its utility, we applied this platform to study the effects of 32 different combinations of five extracellular matrix molecules (collagen I, collagen III, collagen IV, laminin and fibronectin) on cellular differentiation in two contexts: maintenance of primary rat hepatocyte phenotype indicated by intracellular albumin staining and differentiation of mouse embryonic stem (ES) cells toward an early hepatic fate, indicated by expression of a β-galactosidase reporter fused to the fetal liver-specific gene, Ankrd17 (also known as gtar). Using this technique, we identified combinations of ECM that synergistically impacted both hepatocyte function and ES cell differentiation. This versatile technique can be easily adapted to other applications, as it is amenable to studying almost any insoluble microenvironmental cue in a combinatorial fashion and is compatible with several cell types.

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Figure 1: Schematic depiction of fabrication and use of ECM microarray.
Figure 2: Characterization of ECM microarray by indirect immunofluorescence.
Figure 3: Primary rat hepatocytes on ECM microarrays.
Figure 4: Cultured hepatocytes show differential intracellular albumin staining in response to matrix composition.
Figure 5: I114 ES cells differentiate on ECM microarrays.

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Acknowledgements

We would like to thank L. Forrester (University of Edinburgh) for providing the I114 cell line, C. Largent (Cel Associates) and D. Schabacker (Argonne) for useful discussions about acrylamide-gel pad fabrication, T. Martinsky (Telechem) for printing parameters, J. Norwich (S. Chien Lab), R. Agustin (imaging, J. Price Lab), J. Emond and R. Lieber (biostatistics), S. Khetani (cell patterning), J. Felix (hepatocyte isolation) and O. Tran. Funding was generously provided by the US National Institutes of Health, the National Institute of Diabetes and Digestive and Kidney Diseases, the National Science Foundation's Faculty Early Career Development (CAREER) Program and the David and Lucile Packard Foundation.

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  1. Departments of Bioengineering and Medicine, University of California San Diego, 9500 Gilman Drive- MC 0412, La Jolla, 92093-0412, California, USA

    Christopher J Flaim, Shu Chien & Sangeeta N Bhatia

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Correspondence to Sangeeta N Bhatia.

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Flaim, C., Chien, S. & Bhatia, S. An extracellular matrix microarray for probing cellular differentiation. Nat Methods 2, 119–125 (2005). https://doi.org/10.1038/nmeth736

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