Biocompatibility of printed paper-based arrays for 2-D cell cultures Helka Juvonen a,⇑ , Anni Määttänen a , Patrick Laurén b , Petri Ihalainen a , Arto Urtti c , Marjo Yliperttula b , Jouko Peltonen a a Center of Excellence for Functional Materials, Laboratory of Physical Chemistry, Abo Akademi University, Porthaninkatu 3, FI-20500 Turku, Finland b Division of Biopharmaceutics and Pharmacokinetics, Faculty of Pharmacy, University of Helsinki, Finland c Centre for Drug Research, Faculty of Pharmacy, University of Helsinki, Finland article info Article history: Received 29 August 2012 Received in revised form 26 November 2012 Accepted 28 January 2013 Available online 4 February 2013 Keywords: Printed cell arrays Paper substrate ARPE-19 cells Surface energy Roughness abstract The use of paper-based test platforms in cell culture experiments is demonstrated. The arrays used for two-dimensional cell cultures were prepared by printing patterned structures on a paper substrate using a hydrophobic polydimethylsiloxane (PDMS) ink. The non-printed, PDMS-free areas formed the array for the cell growth experiments. Cell imaging was enabled by using a lipophilic staining agent. A set of coated paper substrates was prepared to study the effect of the physicochemical properties of the substrate (topography, roughness and surface energetics) on cell attachment and growth. The studied paper sub- strates were found to be cell-repellent or cell-supporting. Cell growth was supported by substrates with a large bearing area, low surface area ratio (S dr ), high total surface free energy and an intermediate elec- tron donor surface energy component. The cells were grown to full confluency within 72 h. Ó 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved. 1. Introduction There is a growing interest in using paper as a substrate for low- cost analytical tests in healthcare applications. Various printed pa- per-based diagnostic and pharmaceutical applications have re- cently been demonstrated [1–5]. Fields in drug research, chemical toxicity and tissue engineering benefit from the develop- ment of fast cell assays which can be used in high-throughput screening. Several ways of producing platforms for two-dimen- sional (2-D) cell culture have been introduced [6]. Generally, smooth polymer substrates are used as cell culture scaffolds. In or- der to prepare scaffolds with certain topographical features for cell studies, microfabrication methods such as hot embossing [7] and soft lithography [6] have been used, but they are often time con- suming and are not well suited for mass production. Special sur- face treatments, e.g. protein coatings, are also often needed [8]. Fast and cost-efficient methods to produce biocompatible assays can be realized by using roll-to-roll compatible fabrication meth- ods such as printing. Surface properties of the substrate affect cell growth, viability and proliferation [9] and, consequently, cell growth can be gov- erned by surface modification and patterning [8,10,11]. Different cell types respond in different ways to surface topography and roughness of the substrate [11]. Surface energy and its components are important factors that influence the attachment of cells [12– 15]. Surface chemistry has a self-evident impact on surface energy. It also plays a role in specific adhesive binding of cells. Optimal cell adhesion is dependent on the distribution of the beneficial chemi- cal groups (i.e. binding sites) on the surface. Specific interactions of extracellular matrix (ECM) proteins with cells are important for cell growth [9]. The presence of serum proteins in the cell medium and their adhesion to the surface of the substrate have an impor- tant effect on cell attachment [14]. In addition, cell growth can be directed into desired areas by using cell-repellent materials. Polydimethylsiloxane (PDMS) has been proven to hinder the attachment of several cell types due to its low surface energy [16,17]; furthermore, it has been shown to be nontoxic for ARPE- 19 cells [18]. The influence of the characteristics of systematically patterned surfaces on the attachment, spreading and alignment of cells has been investigated at the subcellular level [7,9,19]. Paper, being flexible and porous, offers an interesting alternative to the conven- tional substrates, such as polystyrene and glass, that are often used in cell studies. Uncoated paper has been used in 3-D cell culture applications [20]. The topography and surface chemistry of coated paper substrates can be tailored by changing the composition of the coating formulations and by using different coating methods. Paper can be readily adapted for printing of various functional ink materials [4,21]. For example, the fabrication of printable pla- nar reaction arrays that are applicable for diagnostics has been re- cently demonstrated [4]. In addition, paper is an excellent substrate for the printing of drug substances [2], and can thus be potentially exploited, for example, in high-throughput screening 1742-7061/$ - see front matter Ó 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.actbio.2013.01.033 ⇑ Corresponding author. Tel.: +358 2 2154252; fax: +358 2 2330228. E-mail address: helka.juvonen@abo.fi (H. Juvonen). Acta Biomaterialia 9 (2013) 6704–6710 Contents lists available at SciVerse ScienceDirect Acta Biomaterialia journal homepage: www.elsevier.com/locate/actabiomat