Precise cell patterning using cytophobic self-assembled monolayer deposited on top of semi-transparent gold
First Online: 23 June 2010 DOI:
10.1007/s10544-010-9448-8 Cite this article as: Jing, G., Perry, S.F. & Tatic-Lucic, S. Biomed Microdevices (2010) 12: 935. doi:10.1007/s10544-010-9448-8 Abstract
This paper reports a simple and effective method for cell patterning by using a self-assembled monolayer (SAM)-treated glass surface which is surrounded by semi-transparent gold coated with another type of SAM. Specifically, a hydrophobic SAM, derived from 1-hexadecanethiol (HDT), was coated on the gold surface to prevent cell growth, and a hydrophilic SAM, derived from 3-trimethoxysilyl propyl-diethylenetriamine (DETA), was coated on the exposed glass surface to promote cell growth. The capabilities of this technique are as follows: 1) single-cell resolution, 2) easy alignment of the cell patterns to the structures already existing on the substrate, 3) visualization and verification of the predefined cytophobic/cytophilic pattern prior to cell growth, and 4) convenient monitoring cell growth at the same location for an extended long term period of time. Whereas a number of earlier techniques have demonstrated the single cell resolution, or visualization and verification of the cytophobic/cytophilic patterns prior to cell growth, we believe that our technique is unique in possessing all of these beneficial qualities at the same time. The distinguishing characteristic of our technique is, however, that the use of semi-transparent Cr/Au film allows for convenient brightfield pattern visualization and offers an advantage over previously developed methods which require fluorescent imaging. We have successfully demonstrated the patterning of four different kinds of cells using this technique: immortalized mouse hypothalamic neurons (GT1-7), mouse osteoblast cells (MC3T3), mouse fibroblast cells (NIH3T3) and primary rat hippocampal neurons. This study was performed with a specific ultimate application—the creation of a
multi electrode array (MEA) with predefined localization of cell bodies on top of the electrodes, as well as predefined patterns for cell extensions to grow in between the electrodes. With that goal in mind, we have also determined critical parameters for patterning of each of these cell types, such as the minimum size of a cell-adherent island for exclusively anchoring one cell or two cells, as well as the width of the cytophilic pathway between two islands that enables cell extensions to grow, while preventing the anchoring of the cell bodies. Additionally, we have provided statistical analysis of the occupancy for various sizes and shape of cell-anchoring islands. As demonstrated here, we have developed a novel and reliable cell patterning technique, which can be utilized in various applications, such as biosensors or tissue engineering. Keywords Cell patterning Self-assembled monolayers (SAMs) Single cell patterning References
D.W. Branch, B.C. Wheeler, G.J. Brewer, D.E. Leckband, Long-term maintenance of patterns of hippocampal pyramidal cells onsubstrates of polyethylene glycol and microstamped polylysine. IEEE Trans. Biomed. Eng.
, 290–300 (2000)
G.J. Brewer, J.R. Torricelli, E.K. Evege, P.J. Price, Optimized survival of hippocampal neurons in B 27-supplemented neurobasal, a new serum-free medium combination. J. Neurosci. Res.
, 567–576 (1993)
J.C. Chang, G.J. Brewer, B.C. Wheeler, A modified microstamping technique enhances polylysine transfer and neuronal cell patterning. Biomaterials
, 2863–2870 (2003)
C.S. Chen, M. Mrksich, S. Huang, G.M. Whitesides, D.E. Ingber, Geometric control of cell life and death. Science
, 1425–1428 (1997)
D.T. Chiu, N.L. Jeon, S. Huang, R.S. Kane, C.J. Wargo, I.S. Choi, D.E. Ingber, G.M. Whitesides, Patterned deposition of cells and proteins onto surfaces by using three-dimensional microfluidic systems. Proc. Natl Acad. Sci.
, 2408–2413 (2000)
H.G. Craighead, S.W. Turner, R.C. Davis, C. James, A.M. Perez, P.M. St. John, M.S. Isaacson, L. Kam, W. Shain, J.N. Turner, Chemical and topographical surface modification for control of central nervous system cell adhesion. Biomed. Microdevices
, 49–64 (1998)
H.G. Craighead, C.D. James, A.M.P. Turner, Chemical and topographical patterning for directed cell attachment. Curr. Opin. Solid State Mater. Sci.
, 177–184 (2001)
U. Demirci, G. Montesano, Single cell epitaxy by acoustic picolitre droplets. Lab Chip
, 1139–1145 (2007)
L.E. Dike, C.S. Chen, M. Mrksich, J. Tien, G.M. Whitesides, D.E. Ingber, Geometric control of switching between growth, apoptosis, and differentiation during angiogenesis using micropatterned substrates. In Vitro Cell. Dev. Biol. Anim.
, 441–448 (1999)
D. Falconnet, G. Csucs, H. Michelle Grandin, M. Textor, Surface engineering approaches to micropattern surfaces for cell-based assays. Biomaterials
, 3044–3063 (2006)
D. Gray, J. Tan, J. Voldman, C. Chen, Dielectrophoretic registration of living cells to a microelectrode array. Biosens. Bioelectron.
, 1765–1774 (2004)
H. Gray, O. Fedun, Technical Bulletin# 405—Preparation of BD Falcon™ Cell Culture Inserts for Scanning Electron Microscopy. (2006)
W. He, C. Halberstadt, K. Gonsalves, Lithography application of a novel photoresist for patterning of cells. Biomaterials
, 2055–2063 (2004)
J.J. Hickman, D.A. Stenger, Interactions of Cultured Neurons with Defined Surfaces, in
Enabling Technologies for Cultured Neural Networks, ed. by D.A. Stenger, T.M. McKenna (Academic, San Diego, 1994), pp. 51–76
J. Hu, J. Shi, F. Zhang, L. Lei, X. Li, L. Wang, L. Liu, Y. Chen, High resolution and hybrid patterning for single cell attachment. Microelectron. Eng.
, 726–729 (2010)
B. Ilic, H.G. Craighead, Topographical patterning of chemically sensitive biological materials using a polymer-based dry lift off. Biomed. Microdevices
, 317–322 (2000)
C.D. James, A.J.H. Spence, N.M. Dowell-Mesfin, R.J. Hussain, K.L. Smith, H.G. Craighead, M.S. Isaacson, W. Shain, J.N. Turner, Extracellular recordings from patterned neuronal networks using planar microelectrode arrays. IEEE Trans. Biomed. Eng.
, 1640–1648 (2004)
X. Jiang, D.A. Bruzewicz, A.P. Wong, M. Piel, G.M. Whitesides, Directing cell migration with asymmetric micropatterns. Proc. Natl Acad. Sci.
, 975–978 (2005)
G. Jing, J.P. Labukas, A. Iqbal, S.F. Perry, G.S. Ferguson, S. Tatic-Lucic, A novel method for accurate patterning and positioning of biological cells. Proceedings of SPIE Europe: Microtechnologies for the new millenium, Maspalomas, Grana Canaria, Spain (2007)
G. Jing, Y. Yao, M. Gnerlich, S. Perry, S. Tatic-Lucic, Towards a multi-electrode array (MEA) system for patterned neural networks. Procedia Chem.
, 329–332 (2009)
P.M. St. John, L. Kam, S.W. Turner, H.G. Craighead, M. Issacson, J.N. Turner, W. Shain, Preferential glial cell attachment to microcontact printed surfaces. J. Neurosci. Meth.
, 171–177 (1997)
L. Kam, W. Shain, J.N. Turner, R. Bizios, Correlation of astroglial cell function on micro-patterned surfaces with specific geometric parameters. Biomaterials
, 2343–2350 (1999)
R.S. Kane, S. Takayama, E. Ostuni, D.E. Ingber, G.M. Whitesides, Patterning proteins and cells using soft lithography. Biomaterials
, 2363–2376 (1999)
D. Kleinfeld, K.H. Kahler, P.E. Hockberger, Controlled outgrowth of dissociated neurons on patterned substrates. J. Neurosci.
8, 4098–4120 (1988)
P. Krsko, T. McCann, T. Thach, T. Laabs, H. Geller, M. Libera, Length-scale mediated adhesion and directed growth of neural cells by surface-patterned poly (ethylene glycol) hydrogels. Biomaterials
, 721–729 (2009)
B.M. Lamb, N.P. Westcott, M.N. Yousaf, Live-cell fluorescence microscopy of directed cell migration on partially etched electroactive sam gold surfaces. Chembiochem
, 2220–2224 (2008)
Z. Liposits, I. Merchenthaler, W.C. Wetsel, J.J. Reid, P.L. Mellon, R.I. Weiner, A. Negro-Vilar, Morphological characterization of immortalized hypothalamic neurons synthesizing luteinizing hormone-releasing hormone. Endocrinology
, 1575–1583 (1991)
S. Makohliso, L. Giovangrandi, D. Leonard, H. Mathieu, M. Ilegems, P. Aebischer, Application of Teflon-AF(R) thin films for bio-patterning of neural cell adhesion1. Biosens. Bioelectron.
, 1227–1235 (1998)
R. McBeath, D.M. Pirone, C.M. Nelson, K. Bhadriraju, C.S. Chen, Cell shape, cytoskeletal tension, and rhoa regulate stem cell lineage commitment. Dev. Cell
, 483–495 (2004)
J.S. Miller, M.I. Bethencourt, M. Hahn, T.R. Lee, J.L. West, Laser-scanning lithography (LSL) for the soft lithographic patterning of cell-adhesive self-assembled monolayers. Biotechnol. Bioeng.
, 1060 (2006)
N. Mittal, A. Rosenthal, J. Voldman, nDEP microwells for single-cell patterning in physiological media. Lab Chip
, 1146–1153 (2007)
R.D. Piner, J. Zhu, F. Xu, S. Hong, C.A. Mirkin, Dip-pen nanolithography. Science
, 661–663 (1999)
S. Prasad, X. Zhang, M. Yang, C. Ozkan, M. Ozkan, Neurons as sensors: individual and cascaded chemical sensing. Biosens. Bioelectron.
, 1599–1610 (2004)
E.A. Roth, T. Xu, M. Das, C. Gregory, J.J. Hickman, T. Boland, Inkjet printing for high-throughput cell patterning. Biomaterials
, 3707–3715 (2004)
K. Salaita, Y. Wang, C.A. Mirkin, Applications of dip-pen nanolithography. Nat. Nanotechnol.
, 145–155 (2007)
T. Shibata, S. Yamanaka, N. Kato, T. Kawashima, M. Nomura, T. Mineta, E. Makino, Fabrication of micromanipulator array for cell patterning. Microelectron. Eng.
, 1439–1442 (2009)
R.K. Smith, P.A. Lewis, P.S. Weiss, Patterning self-assembled monolayers. Prog. Surf. Sci.
, 1–68 (2004)
N. Sniadecki, A. Anguelouch, M. Yang, C. Lamb, Z. Liu, S. Kirschner, Y. Liu, D. Reich, C. Chen, Magnetic microposts as an approach to apply forces to living cells. Proc. Natl Acad. Sci.
, 14553 (2007)
H. Thissen, G. Johnson, P. Hartley, P. Kingshott, H. Griesser, Two-dimensional patterning of thin coatings for the control of tissue outgrowth. Biomaterials
, 35–43 (2006)
T. Tzvetkova-Chevolleau, E. Yoxall, D. Fuard, F. Bruckert, P. Schiavone, M. Weidenhaupt, Microscale adhesion patterns for the precise localization of amoeba. Microelectron. Eng.
, 1485–1487 (2009)
G. Vunjak-Novakovic, Patterning stem cell differentiation. Cell Stem Cell
, 362–363 (2008)
G.M. Whitesides, E. Ostuni, S. Takayama, X. Jiang, D.E. Ingber, Soft lithography in biology and biochemistry. Annu. Rev. Biomed. Eng.
, 335–373 (2001)
Y. Xia, G.M. Whitesides, Soft lithography. Angew. Chem. Int. Ed.
, 550–575 (1998)
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