Arrayed cellular microenvironments for identifying culture and differentiation conditions for stem, primary and rare cell populations

Arrayed cellular microenvironments for identifying culture and differentiation conditions for stem, primary and rare cell populations

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ABSTRACT During the development of an organism, cells are exposed to a myriad of signals, structural components and scaffolds, which collectively make up the cellular microenvironment. The


majority of current developmental biology studies examine the effect of individual or small subsets of molecules and parameters on cellular behavior, and they consequently fail to explore


the complexity of factors to which cells are exposed. Here we describe a technology, referred to as arrayed cellular microenvironments (ACMEs), that allows for a high-throughput examination


of the effects of multiple extracellular components in a combinatorial manner on any cell type of interest. We will specifically focus on the application of this technology to human


pluripotent stem cells (hPSCs), a population of cells with tremendous therapeutic potential, and one for which growth and differentiation conditions are poorly characterized and far from


defined and optimized. A standard ACME screen uses the technologies previously applied to the manufacture and analysis of DNA microarrays, requires standard cell-culture facilities and can


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supported by funding from the University of California San Diego Stem Cell Program and by a gift from Michael and Nancy Kaehr. This research was supported in part by the California Institute


of Regenerative Medicine (RS1-00172-1) to S.C. and (RB1-01406) to K.W. AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Cellular and Molecular Medicine, Stem Cell Program, University of


California, San Diego, California, USA David A Brafman & Karl Willert * Department of Bioengineering, University of California, San Diego, California, USA Shu Chien Authors * David A


Brafman View author publications You can also search for this author inPubMed Google Scholar * Shu Chien View author publications You can also search for this author inPubMed Google Scholar


* Karl Willert View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS D.A.B., S.C. and K.W. developed the protocol. D.A.B. and K.W. designed and


performed the experiments. D.A.B., S.C. and K.W. analyzed the results. D.A.B., S.C. and K.W. wrote the manuscript. CORRESPONDING AUTHORS Correspondence to David A Brafman or Karl Willert.


ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing financial interests. SUPPLEMENTARY INFORMATION SUPPLEMENTARY FIG 1 Schematic representation of ACME format. (a)


Schematic of a 10x10 subarray that contains 20 different conditions spotted in replicates of five. Spot diameter (closed circles) is 150 μm and the center to center distance between


neighboring spots is 450 μm. These subarrays can be arranged to generate several different designs including (b) a 8x2, 1600 spot/320 condition, (c) a 16x4, 6400 spot/1280 condition, or (d)


a 16x5, 8000 spot/1600 condition array. (PPT 415 kb) SUPPLEMENTARY TABLE 1 Example of analysis of raw data generated from the array in Figure 5e. (DOC 1214 kb) RIGHTS AND PERMISSIONS


Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Brafman, D., Chien, S. & Willert, K. Arrayed cellular microenvironments for identifying culture and differentiation


conditions for stem, primary and rare cell populations. _Nat Protoc_ 7, 703–717 (2012). https://doi.org/10.1038/nprot.2012.017 Download citation * Published: 15 March 2012 * Issue Date:


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