Microarray technology has become extremely useful in expediting the investigation of large libraries of materials in a variety of biomedical applications, such as in DNA chips, protein and cellular microarrays. In the development of cellular microarrays, traditional high-throughput printing strategies on stiff, glass substrates and non-covalent attachment methods are limiting. We have developed a facile strategy to fabricate multifunctional high-throughput microarrays embedded at the surface of a hydrogel substrate using thiol-ene chemistry. This user-friendly method provides a platform for the immobilization of a combination of bioactive and diagnostic molecules, such as peptides and dyes, at the surface of poly(ethylene glycol)-based hydrogels. The robust and orthogonal nature of thiol-ene chemistry allows for a range of covalent attachment strategies in a fast and reliable manner, and two complementary strategies for the attachment of active molecules are demonstrated.
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Castel, D., Pitaval, A., Debily, M. & Gidrol, X. Cell microarrays in drug discovery. Drug Discov. Today 11, 616–622 (2006).
Schulze, A. & Downward, J. Navigating gene expression using microarrays – a technology review. Nature Cell Biol. 3, 190–195 (2001).
Ramsay, G. DNA chips: state-of-the-art. Nature Biotechnol. 16, 40–44 (1998).
Anderson, D. et al. Biomaterial microarrays: rapid, microscale screening of polymer–cell interaction. Biomaterials 26, 4892–4897 (2005).
Yliperttula, M. et al. High-throughput screening of cell responses to biomaterials. Eur. J. Pharm. Sci. 35, 151–160 (2008).
Albrecht, D., Tsang, V., Sah, R. & Bhatia, S. Photo- and electropatterning of hydrogel-encapsulated living cell arrays. Lab Chip 5, 111–118 (2005).
Yamada, K. M. Adhesive recognition sequences. J. Biol. Chem. 266, 12809–12812 (1991).
Hoffman, L. & Carpenter, M. Characterization and culture of human embryonic stem cells. Nature Biotechnol. 23, 699–708 (2005).
Evenram, S., Artym, V. & Yamada, K. Matrix control of stem cell fate. Cell 126, 645–647 (2006).
Li, Y. et al. Hydrogels as artificial matrices for human embryonic stem cell self-renewal. J. Biomed. Mater. Res. A 79A, 1–5 (2006).
Mohr, J., Depablo, J. & Palecek, S. 3-D microwell culture of human embryonic stem cells. Biomaterials 27, 6032–6042 (2006).
Moeller, H. et al. A microwell array system for stem cell culture. Biomaterials 29, 752–763 (2008).
Yeo, D. S. Y., Panicker, R. C., Tan, L. P. & Yao, S. Q. Strategies for immobilization of biomolecules in a microarray. Comb. Chem. High Throughput Screening 7, 213–221 (2004).
Houseman, B. T., Huh, J. H., Kron, S. J. & Mrksich, M. Peptide chips for the quantitative evaluation of protein kinase activity. Nature Biotechnol. 20, 270–274 (2002).
Köhn, M. et al. Staudinger ligation: a new immobilization strategy for the preparation of small-molecule arrays. Angew. Chem. Int. Ed. 42, 5830–5834 (2003).
Falsey, J. R. et al. Peptide and small molecule microarray for high throughput cell adhesion and functional assays. Bioconjugate Chem. 12, 346–353 (2001).
Christman, K. L., Broyer, R. M., Tolstyka, Z. P. & Maynard, H. D. Site-specific protein immobilization through N-terminal oxime linkages. J. Mater. Chem. 17, 2021–2027 (2007).
Fukuda, J. et al. Micropatterned cell co-cultures using layer-by-layer deposition of extracellular matrix components. Biomaterials 27, 1479–1486 (2006).
Derda, R. et al. Defined substrates for human embryonic stem cell growth identified from surface arrays. ACS Chem. Biol. 2, 347–355 (2007).
Northen, T., Greving, M. & Woodbury, N. Combinatorial screening of biomimetic protein affinity materials. Adv. Mater. 20, 4691–4697 (2008).
Tweedie, C., Anderson, D., Langer, R. & Van Vliet, K. Combinatorial material mechanics: high-throughput polymer synthesis and nanomechanical screening. Adv. Mater. 17, 2599–2604 (2005).
Anderson, D., Lynn, D. & Langer, R. Semi-automated synthesis and screening of a large library of degradable cationic polymers for gene delivery. Angew. Chem. Int. Ed. 42, 3153–3158 (2003).
Flaim, C., Chien, S. & Bhatia, S. An extracellular matrix microarray for probing cellular differentiation. Nature Methods 2, 119–125 (2005).
Anderson, D., Levenberg, S. & Langer, R. Nanoliter-scale synthesis of arrayed biomaterials and application to human embryonic stem cells. Nature Biotechnol. 22, 863–866 (2004).
Tourniaire, G. et al. Polymer microarrays for cellular adhesion. Chem. Commun. 2118 (2006).
Stadler, V. et al. Combinatorial synthesis of peptide arrays with a laser printer. Angew. Chem. Int. Ed. 47, 7132–7135 (2008).
Hsiong, S. et al. Differentiation stage alters matrix control of stem cells. J. Biomed. Mater. Res. A 85A, 145–156 (2008).
Saha, K. et al. Biomimetic interfacial interpenetrating polymer networks control neural stem cell behavior. J. Biomed. Mater. Res. A 81A, 240–249 (2007).
Heyries, K. A., Blum, L. J. & Marquette, C. A. Direct poly(dimethylsiloxane) surface functionalization with vinyl modified DNA. Chem. Mater. 20, 1251–1253 (2008).
Kolb, H. C., Finn, M. G. & Sharpless, K. B. Click chemistry: diverse chemical function from a few good reactions. Angew. Chem. Int. Ed. 40, 2004–2021 (2001).
Campos, L. et al. Highly versatile and robust materials for soft imprint lithography based on thiol-ene click chemistry. Adv. Mater. 20, 3728–3733 (2008).
Cushing, M. C. & Anseth, K. Hydrogel cell cultures. Science 316, 1133–1134 (2007).
Hoyle, C., Lee, T. & Roper, T. Thiol-enes: chemistry of the past with promise for the future. J. Polym. Sci., Part A: Polym. Chem. 42, 5301–5338 (2004).
Killops, K. L., Campos, L. M. & Hawker, C. J. Robust, efficient, and orthogonal synthesis of dendrimers via thiol-ene ‘click’ chemistry. J. Am. Chem. Soc. 130, 5062–5064 (2008).
Campos, L. M. et al. Development of thermal and photochemical strategies for thiol-ene click polymer functionalization. Macromolecules 41, 7063–7070 (2008).
Rydholm, A., Reddy, S., Anseth, K. & Bowman, C. Development and characterization of degradable thiol-allyl ether photopolymers. Polymer 48, 4589–4600 (2007).
Polizzotti, B. D., Fairbanks, B. D. & Anseth, K. Three-dimensional biochemical patterning of click-based composite hydrogels via thiolene photopolymerization. Biomacromolecules 9, 1084–1087 (2008).
Ruoslahti, E. RGD and other recognition sequences for integrins. Annu. Rev. Cell Dev. Biol. 12, 697–715 (1996).
Pierschbacher, M. D. & Ruoslahti, E. Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature 309, 30–33 (1984).
Dawson, P. E., Churchill, M. J., Ghadiri, M. R. & Kent, S. B. H. Modulation of reactivity in native chemical ligation through the use of thiol additives. J. Am. Chem. Soc. 119, 4325–4329 (1997).
Shao, J. & Tam, J. P. Unprotected peptides as building-blocks for the synthesis of peptide dendrimers with oxime, hydrazone and thiazolidine linkages. J. Am. Chem. Soc. 117, 3893–3899 (1995).
Dedola, S., Nepogodiev, S. & Field, R. Recent applications of the Cu(i)-catalysed Huisgen azide-alkyne 1,3-dipolar cycloaddition reaction in carbohydrate chemistry. Org. Biomol. Chem. 5, 1006–1017 (2007).
Beatty, K. E., Xie, F., Wang, Q. & Tirrell, D. A. Selective dye-labeling of newly synthesized proteins in bacterial cells. J. Am. Chem. Soc. 127, 14150–14151 (2005).
This work made use of Materials Research Laboratory's Central Facilities supported by the Materials Research Science & Engineering Centers Program of the National Science Foundation (No. DMR05-20415). N.G. thanks the National Science Foundation for a Graduate Research Fellowship. L.M.C. thanks the University of California for a Presidential Fellowship. We gratefully acknowledge the use of the UCSB Laboratory for Stem Cell Biology and Engineering and funding from the California Institute for Regenerative Medicine.
The authors declare no competing financial interests.
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