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Lithographically Defined Two- and Three-Dimensional Tissue Microarrays

  • Esther W. Gomez
  • Celeste M. Nelson
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 671)

Abstract

Traditional methods to study normal and pathological development of tissues have been limited by ­difficulties in controlling experimental conditions and quantifying biological processes of interest. Here we describe methods to create microarrays of engineered tissues that enable controlled and quantitative investigations. Using soft lithography-based techniques, extracellular matrix proteins can be microcontact printed or micromolded to make two- and three-dimensional micropatterned scaffolds. The ultimate form and resulting properties of the tissue construct are dictated by the geometry of the patterned extracellular matrix components. This chapter describes elastomeric stamp fabrication, microcontact printing and micromolding of extracellular matrix proteins, cell culture in micropatterned substrata, and quantitative immunofluorescence analysis of micropatterned tissues.

Key words

Tissue engineering Microfabrication Organotypic culture Epithelial 

Notes

Acknowledgments

This work was supported by grants from the NIH (CA128660 and GM083997), Susan G. Komen for the Cure (FAS0703855); and the David & Lucile Packard Foundation. C.M.N. holds a Career Award at the Scientific Interface from the Burroughs Wellcome Fund. E.W.G. was supported by post doctoral fellowships from the New Jersey Commission on Cancer Research and Susan G. Komen for the cure.

References

  1. 1.
    Langer, R., and Vacanti, J. P. (1993) Tissue engineering. Science 260, 920–926.CrossRefGoogle Scholar
  2. 2.
    O’Neill, C., Jordan, P., and Ireland, G. (1986) Evidence for two distinct mechanisms of anchorage stimulation in freshly explanted and 3t3 swiss mouse fibroblasts. Cell 44, 489–496.CrossRefGoogle Scholar
  3. 3.
    Chen, C. S., Mrksich, M., Huang, S., Whitesides, G. M., and Ingber, D. E. (1997) Geometric control of cell life and death. Science 276, 1425–1428.CrossRefGoogle Scholar
  4. 4.
    Whitesides, G. M., Ostuni, E., Takayama, S., Jiang, X., and Ingber, D. E. (2001) Soft lithography in biology and biochemistry. Annu. Rev. Biomed. Eng. 3, 335–373.CrossRefGoogle Scholar
  5. 5.
    Nelson, C. M., Jean, R. P., Tan, J. L., Liu, W. F., Sniadecki, N. J., Spector, A. A., and Chen, C. S. (2005) Emergent patterns of growth controlled by multicellular form and mechanics. Proc. Natl. Acad. Sci. USA 102, 11594–11599.CrossRefGoogle Scholar
  6. 6.
    Tien, J., Nelson, C. M., and Chen, C. S. (2002) Fabrication of aligned microstructures with a single elastomeric stamp. Proc. Natl. Acad. Sci. USA 99, 1758–1762.CrossRefGoogle Scholar
  7. 7.
    Jeon, N. L., Dertinger, S. K. W., Chiu, D. T., Choi, I. S., Stroock, A. D., and Whitesides, G. M. (2000) Generation of solution and surface gradients using microfluidic systems. Langmuir 16, 8311–8316.CrossRefGoogle Scholar
  8. 8.
    Ostuni, E., Kane, R., Chen, C. S., Ingber, D. E., and Whitesides, G. M. (2000) Patterning mammalian cells using elastomeric membranes. Langmuir 16, 7811–7819.CrossRefGoogle Scholar
  9. 9.
    Flaim, C. J., Chien, S., and Bhatia, S. N. (2005) An extracellular matrix microarray for probing cellular differentiation. Nat. Methods 2, 119–125.CrossRefGoogle Scholar
  10. 10.
    Wilson, D. L., Martin, R., Hong, S., Cronin-Golomb, M., Mirkin, C. A., and Kaplan, D. L. (2001) Surface organization and nanopatterning of collagen by dip-pen nanolithography. Proc. Natl. Acad. Sci. USA 98, 13660–13664.CrossRefGoogle Scholar
  11. 11.
    Jackson, B. L., and Groves, J. T. (2007) Hybrid protein-lipid patterns from aluminum templates. Langmuir 23, 2052–2057.CrossRefGoogle Scholar
  12. 12.
    Nguyen, K. T., and West, J. L. (2002) Photopolymerizable hydrogels for tissue engineering applications. Biomaterials 23, 4307–4314.CrossRefGoogle Scholar
  13. 13.
    Tsang, V. L., Chen, A. A., Cho, L. M., Jadin, K. D., Sah, R. L., DeLong, S., West, J. L., and Bhatia, S. N. (2007) Fabrication of 3d hepatic tissues by additive photopatterning of cellular hydrogels. Faseb J. 21, 790–801.CrossRefGoogle Scholar
  14. 14.
    Tang, M. D., Golden, A. P., and Tien, J. (2003) Molding of three-dimensional microstructures of gels. J. Am. Chem. Soc. 125, 12988–12989.CrossRefGoogle Scholar
  15. 15.
    Tang, M. D., Golden, A. P., and Tien, J. (2004) Fabrication of collagen gels that contain patterned, micrometer-scale cavities. Adv. Mater. 16, 1345–1348.CrossRefGoogle Scholar
  16. 16.
    Nelson, C. M., Vanduijn, M. M., Inman, J. L., Fletcher, D. A., and Bissell, M. J. (2006) Tissue geometry determines sites of mammary branching morphogenesis in organotypic cultures. Science 314, 298–300.CrossRefGoogle Scholar
  17. 17.
    Fukuda, J., Khademhosseini, A., Yeo, Y., Yang, X. Y., Yeh, J., Eng, G., Blumling, J., Wang, C. F., Kohane, D. S., and Langer, R. (2006) Micromolding of photocrosslinkable chitosan hydrogel for spheroid microarray and co-cultures. Biomaterials 27, 5259–5267.CrossRefGoogle Scholar
  18. 18.
    Nelson, C. M., Inman, J. L., and Bissell, M. J. (2008) Three-dimensional lithographically defined organotypic tissue arrays for quantitative analysis of morphogenesis and neoplastic progression. Nat. Protoc. 3, 674–678.CrossRefGoogle Scholar
  19. 19.
    Jongpaiboonkit, L., King, W. J., Lyons, G. E., Paguirigan, A. L., Warrick, J. W., Beebe, D. J., and Murphy, W. L. (2008) An adaptable hydrogel array format for 3-dimensional cell culture and analysis. Biomaterials 29, 3346–3356.CrossRefGoogle Scholar
  20. 20.
    Fukuda, J., Khademhosseini, A., Yeh, J., Eng, G., Cheng, J. J., Farokhzad, O. C., and Langer, R. (2006) Micropatterned cell co-cultures using layer-by-layer deposition of extracellular matrix components. Biomaterials 27, 1479–1486.CrossRefGoogle Scholar
  21. 21.
    Bettinger, C. J., Weinberg, E. J., Kulig, K. M., Vacanti, J. P., Wang, Y. D., Borenstein, J. T., and Langer, R. (2006) Three-dimensional microfluidic tissue-engineering scaffolds using a flexible biodegradable polymer. Adv. Mater. 18, 165–169.CrossRefGoogle Scholar
  22. 22.
    Golden, A. P., and Tien, J. (2007) Fabrication of microfluidic hydrogels using molded gelatin as a sacrificial element. Lab Chip 7, 720–725.CrossRefGoogle Scholar
  23. 23.
    Tan, J. L., Liu, W., Nelson, C. M., Raghavan, S., and Chen, C. S. (2004) Simple approach to micropattern cells on common culture substrates by tuning substrate wettability. Tissue Eng. 10, 865–872.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Esther W. Gomez
    • 1
  • Celeste M. Nelson
    • 2
  1. 1.Departments of Chemical Engineering and Molecular BiologyPrinceton UniversityPrincetonUSA
  2. 2.Department of Chemical EngineeringPrinceton UniversityPrincetonUSA

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