Biomaterials 25 (2004) 4073–4078 Ordered growth of neurons on diamond Christian G. Specht a,1 , Oliver A. Williams b,1 , Richard B. Jackman b, *, Ralf Schoepfer a, * a Laboratory for Molecular Pharmacology, Department of Pharmacology, UCL, Gower Street, London WC1E 6BT, UK b Diamond Electronics Group, Department of Electronic and Electrical Engineering, UCL, Gower Street, London WC1E 6BT, UK Received 22 September 2003; accepted 8 November 2003 Abstract Diamond has a number of unique properties that make it an attractive electronic and bio-electronic material. Here we show the ordered growth of mammalian neurons, the principal electrogenic cells of the nervous system, on diamond. Proteins were specifically patterned on diamond surfaces by micro-contact printing. Mouse cortical neurons were then cultured on these substrates. Neuron adhesion and outgrowth was specific for those areas of the diamond that had been stamped with laminin, resulting in ordered growth of high resolution. Neurons survived in culture for the duration of the experiment, and laminin patterns were stable for at least 1 week in culture. The relative biocompatibility of diamond and the suitability of neuron interfacing with the hydrogen surface conductivity layer make this an interesting model for the formation of defined neuronal networks and for implants. r 2004 Elsevier Ltd. All rights reserved. Keywords: Biocompatibility; Diamond; Laminin; Micro-contact printing (mCP); Network; Neuron 1. Introduction Diamond, the tetrahedral allotrope of carbon, has a number of unusual properties that make it attractive as a biomaterial. Its extreme chemical inertness, for example, makes it highly resistant to wet etching or other forms of degradation [1]. Diamond generally shows excellent biocompatibility [2,3]. Much of the work to date has been performed on diamond-like carbon (DLC), an amorphous concoction of both sp 2 and sp 3 phases, that are ideal for coating materials used for prosthesis [4,5]. However, the work here focuses on single crystal diamond, i.e. 100% sp 3 carbon. Single crystal diamond can be grown as an atomically flat surface [6]. It is optically transparent and functions as a wide band-gap semiconductor. The diamond surface can be made hydrophobic or hydrophilic by hydrogen or oxygen termination, respectively [7,8]. The hydrogen-terminated surface exhibits a unique p-type surface conductivity that may be used for device fabrication [9], and highly sensitive ion and pH sensors have been demonstrated on this layer [10,11]. The surface termination of diamond, hence the respective surface conductivity layer, can also be patterned at the nanoscale [12]. This approach is an interesting alter- native to self-assembled monolayers (SAMS) for creat- ing localised hydrophobic/hydrophilic regions with the simple reversal of surface termination, and has been used to fabricate a single-hole transistor [13]. Furthermore, diamond surfaces are extremely stable, irrespective of the termination (with hydrogen or oxygen) [14]. Recently, it has been shown that the hydrogen-terminated surface is an ideal starting point for the covalent attachment of biomolecules such as DNA [15]. The resulting DNA-functionalised surface could be repeatedly hybridised with labelled oligonu- cleotides without any loss of the fluorescence signal, indicating high stability of the chemical crosslinking. Micro-contact printing (mCP) has been promoted over the past decade as a technique for selectively transfer- ring molecules, such as laminin onto surfaces. Ordered neuron growth on such patterns have been achieved on glass and other materials before [16,17]. To explore the unique properties of diamond mentioned above, we investigated possibilities of ordered neuron growth on this biomaterial. This was achieved by combining mCP, ARTICLE IN PRESS *Corresponding authors. Tel.: +44-20-7679-1382; fax: +44-20- 7388-9325 (R.B. Jackman), Tel.: +44-20-7679-7242; fax: +44-20- 7679-7245 (R. Schoepfer). E-mail addresses: r.jackman@ee.ucl.ac.uk (R.B. Jackman), r.schoepfer@ucl.ac.uk (R. Schoepfer). 1 OAW and CGS contributed equally to this work. 0142-9612/$-see front matter r 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.biomaterials.2003.11.006