Fabrication of a multiplexed artificial cellular microenvironment array

Yasumasa Mashimo, Momoko Yoshioka, Yumie Tokunaga, Christopher Fockenberg, Shiho Terada, Yoshie Koyama, Teiko Shibata-Seki, Koki Yoshimoto, Risako Sakai, Hayase Hakariya, Li Liu, Toshihiro Akaike, Eiry Kobatake, Siew Eng How, Motonari Uesugi, Yong Chen, Ken Ichiro Kamei

Research output: Contribution to journalArticlepeer-review


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.

Original languageEnglish (US)
Article numbere57377
JournalJournal of Visualized Experiments
Issue number139
StatePublished - Sep 7 2018


  • Bioengineering
  • Cellular microenvironment
  • Embryonic stem cell
  • Issue 139
  • Microfluidic
  • Nanofiber
  • Self-renewal
  • Single-cell profiling

ASJC Scopus subject areas

  • General Neuroscience
  • General Chemical Engineering
  • General Biochemistry, Genetics and Molecular Biology
  • General Immunology and Microbiology


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