Affiliations 

  • 1 Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; Department of Life Science and Technology, School of Life Science and Technology, Tokyo Institute of Technology
  • 2 Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University
  • 3 Department of Life Science and Technology, School of Life Science and Technology, Tokyo Institute of Technology
  • 4 Biomaterials Center for Regenerative Medical Engineering, Foundation for Advancement of International Science
  • 5 Faculty of Science and Natural Resources, Universiti Malaysia Sabah
  • 6 Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; Institute for Chemical Research, Kyoto University
  • 7 Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; Ecole Normale Supérieure
  • 8 Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; kkamei@icems.kyoto-u.ac.jp
J Vis Exp, 2018 09 07.
PMID: 30247461 DOI: 10.3791/57377

Abstract

Cellular microenvironments consist of a variety of cues, such as growth factors, extracellular matrices, and intercellular interactions. These cues are well orchestrated and are crucial in regulating cell functions in a living system. Although a number of researchers have attempted to investigate the correlation between environmental factors and desired cellular functions, much remains unknown. This is largely due to the lack of a proper methodology to mimic such environmental cues in vitro, and simultaneously test different environmental cues on cells. Here, we report an integrated platform of microfluidic channels and a nanofiber array, followed by high-content single-cell analysis, to examine stem cell phenotypes altered by distinct environmental factors. To demonstrate the application of this platform, this study focuses on the phenotypes of self-renewing human pluripotent stem cells (hPSCs). Here, we present the preparation procedures for a nanofiber array and the microfluidic structure in the fabrication of a Multiplexed Artificial Cellular MicroEnvironment (MACME) array. Moreover, overall steps of the single-cell profiling, cell staining with multiple fluorescent markers, multiple fluorescence imaging, and statistical analyses, are described.

* Title and MeSH Headings from MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.