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Three-Dimensional Cell-Based Microarrays: Printing Pluripotent Stem Cells into 3D Microenvironments

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1771))

Abstract

Cell-based microarrays are valuable platforms for the study of cytotoxicity and cellular microenvironment because they enable high-throughput screening of large sets of conditions at reduced reagent consumption. However, most of the described microarray technologies have been applied to two-dimensional cultures, which do not accurately emulate the in vivo three-dimensional (3D) cell–cell and cell–extracellular matrix interactions.

Herein, we describe the methodology for production of alginate- and Matrigel-based 3-D cell microarrays for the study of mouse and human pluripotent stem cells on two different chip-based platforms. We further provide protocols for on-chip proliferation/viability analysis and the assessment of protein expression by immunofluorescence.

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References

  1. Azarin SM, Larson EA, Almodóvar-Cruz JM et al (2012) Effects of 3D microwell culture on growth kinetics and metabolism of human embryonic stem cells. Biotechnol Appl Biochem 59:88–96

    Article  CAS  Google Scholar 

  2. Loessner D, Stok KS, Lutolf MP et al (2010) Bioengineered 3D platform to explore cell–ECM interactions and drug resistance of epithelial ovarian cancer cells. Biomaterials 31:8494–8506

    Article  CAS  Google Scholar 

  3. Bruce A, Evans R, Mezan R et al (2015) Three-dimensional microfluidic tri-culture model of the bone marrow microenvironment for study of acute lymphoblastic leukemia. PLoS One 10:e0140506

    Article  Google Scholar 

  4. Eglen RM, Randle DH (2015) Drug discovery goes three-dimensional: goodbye to flat high-throughput screening? Assay Drug Dev Technol 13:262–265

    Article  CAS  Google Scholar 

  5. Rothbauer M, Charwat V, Ertl P (2016) Cell microarrays for biomedical applications. In: Li PC, Sedighi A, Wang L (eds) Microarray technology. Springer, New York, pp 273–291

    Chapter  Google Scholar 

  6. Fernandes TG, Diogo MM, Clark DS et al (2009) High-throughput cellular microarray platforms: applications in drug discovery, toxicology and stem cell research. Trends Biotechnol 27:342–349

    Article  CAS  Google Scholar 

  7. Meli L, Barbosa HSC, Hickey AM et al (2014) Three dimensional cellular microarray platform for human neural stem cell differentiation and toxicology. Stem Cell Res 13:36–47

    Article  CAS  Google Scholar 

  8. Kwon SJ, Lee DW, Shah DA et al (2014) High-throughput and combinatorial gene expression on a chip for metabolism-induced toxicology screening. Nat Commun 5:3739

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Fernandes TG, Kwon S-J, Lee M-Y et al (2008) On-chip, cell-based microarray immunofluorescence assay for high-throughput analysis of target proteins. Anal Chem 80:6633–6639

    Article  CAS  Google Scholar 

  10. Nierode G, Kwon PS, Dordick JS et al (2015) Cell-based assay design for high-content screening of drug candidates. J Microbiol Biotechnol 26:213–225

    Article  Google Scholar 

  11. Joshi P, Lee MY (2015) High content imaging (HCI) on miniaturized three-dimensional (3D) cell cultures. Biosensors 5:768–790

    Article  CAS  Google Scholar 

  12. Thomson JA (1998) Embryonic stem cell lines derived from human blastocysts. Science 282:1145–1147

    Article  CAS  Google Scholar 

  13. Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–676

    Article  CAS  Google Scholar 

  14. Mitalipov S, Wolf D (2009) Totipotency, pluripotency and nuclear reprogramming. In: Engineering of stem cells. Springer Berlin Heidelberg, Berlin, Heidelberg, pp 185–199

    Chapter  Google Scholar 

  15. Klimanskaya I, Rosenthal N, Lanza R (2008) Derive and conquer: sourcing and differentiating stem cells for therapeutic applications. Nat Rev Drug Discov 7:131–142

    Article  CAS  Google Scholar 

  16. Lee M-Y, Kumar RA, Sukumaran SM et al (2008) Three-dimensional cellular microarray for high-throughput toxicology assays. Proc Natl Acad Sci U S A 105:59–63

    Article  CAS  Google Scholar 

  17. Fernandes TG, Kwon S-J, Bale SS et al (2010) Three-dimensional cell culture microarray for high-throughput studies of stem cell fate. Biotechnol Bioeng 106:106–118

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Diogo MM, Henrique D, Cabral JMS (2008) Optimization and integration of expansion and neural commitment of mouse embryonic stem cells. Biotechnol Appl Biochem 49:105

    Article  CAS  Google Scholar 

  19. Lee M-Y, Park CB, Dordick JS et al (2005) Metabolizing enzyme toxicology assay chip (MetaChip) for high-throughput microscale toxicity analyses. Proc Natl Acad Sci U S A 102:983–987

    Article  CAS  Google Scholar 

  20. Nierode GJ, Perea BC, McFarland SK, Pascoal JF, Clark DS, Schaffer DV, Dordick JS (2016) High-throughput toxicity and phenotypic screening of 3D human neural progenitor cell cultures on a microarray chip platform. Stem Cell Rep 7(5):970–982

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The work involved in the development of this protocol was funded by NIH (ES020903) and NYSTEM (C026717). JFP acknowledges support from Fundação para a Ciência e Tecnologia (SFRH/BD/79590/2011), which also funded iBB (UID/BIO/04565/2013).

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Correspondence to Tiago G. Fernandes or Jonathan S. Dordick .

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Pascoal, J., Fernandes, T.G., Nierode, G.J., Diogo, M.M., Dordick, J.S., Cabral, J.M.S. (2018). Three-Dimensional Cell-Based Microarrays: Printing Pluripotent Stem Cells into 3D Microenvironments. In: Ertl, P., Rothbauer, M. (eds) Cell-Based Microarrays. Methods in Molecular Biology, vol 1771. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7792-5_6

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  • DOI: https://doi.org/10.1007/978-1-4939-7792-5_6

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7791-8

  • Online ISBN: 978-1-4939-7792-5

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