Name: polymer microarrays for high throughput discovery of biomaterials
Uploaded: 2012-01-25T13:00:00
Duration: 13 min 37 s
Description: Watch this Scientific Journal Video about Polymer Microarrays for High Throughput Discovery of Biomaterials at JoVE.com

Funding from the Wellcome Trust is kindly acknowledged (grant number 085245/Z/08/Z). Nottingham Nanotechnology and Nanoscience Centre is kindly acknowledged for giving access to the Raman system and for the East Midlands Development Agency for funding this equipment.

<ol>
	<li>Hook, A. L., Anderson, D. G., Langer, R., Williams, P., Davies, M. C., Alexander, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+throughput+methods+applied+in+biomaterial+development+and+discovery.">High throughput methods applied in biomaterial development and discovery.</a> Biomaterials. 31, 187-198 (2010).</li><li>Anderson, D. G., Levenberg, S., Langer, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoliter-scale+synthesis+of+arrayed+biomaterials+and+application+to+human+embryonic+stem+cells.">Nanoliter-scale synthesis of arrayed biomaterials and application to human embryonic stem cells.</a> Nature Biotechnology. 22, 863-866 (2004).</li><li>Mei, Y., Saha, K., Bogatyrev, S. R., Yang, J., Hook, A. L., Kalcioglu, Z. I., Cho, S. W., Mitalipova, M., Pyzocha, N., Rojas, F., Van Vliet, K. J., Davies, M. C., Alexander, M. R., Langer, R., Jaenisch, R., Anderson, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Combinatorial+development+of+biomaterials+for+clonal+growth+of+human+pluripotent+stem+cells.">Combinatorial development of biomaterials for clonal growth of human pluripotent stem cells.</a> Nature Materials. 9, 768-778 (2010).</li><li>Pernagallo, S., Wu, M., Gallagher, M. P., Bradley, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colonising+new+frontiers-microarrays+reveal+biofilm+modulating+polymers.">Colonising new frontiers-microarrays reveal biofilm modulating polymers.</a> Journal of Materials Chemistry. 21, 96-101 (2011).</li><li>Yang, J., Mei, Y., Hook, A. L., Taylor, M., Urquhart, A. J., Bogatyrev, S. R., Langer, R., Anderson, D. G., Davies, M. C., Alexander, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polymer+surface+functionalities+that+control+human+embryoid+body+cell+adhesion+revealed+by+high+throughput+surface+characterization+of+combinatorial+material+microarrays.">Polymer surface functionalities that control human embryoid body cell adhesion revealed by high throughput surface characterization of combinatorial material microarrays.</a> Biomaterials. 31, 8827-8838 (2010).</li><li>Urquhart, A. J., Anderson, D. G., Taylor, M., Alexander, M. R., Langer, R., Davies, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+throughput+surface+characterisation+of+a+combinatorial+material+library.">High throughput surface characterisation of a combinatorial material library.</a> Advanced Materials. 19, 2486-2491 (2007).</li><li>Taylor, M., Urquhart, A. J., Zelzer, M., Davies, M. C., Alexander, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Picoliter+water+contact+angle+measurement+on+polymers.">Picoliter water contact angle measurement on polymers.</a> Langmuir. 23, 6875-6878 (2007).</li><li>Urquhart, A. J., Taylor, M., Anderson, D. G., Langer, R., Davies, M. C., Alexander, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TOF-SIMS+analysis+of+a+576+micropatterned+copolymer+array+to+reveal+surface+moieties+that+control+wettability.">TOF-SIMS analysis of a 576 micropatterned copolymer array to reveal surface moieties that control wettability.</a> Analytical Chemistry. 80, 135-142 (2008).</li><li>Hook, A. L., Yang, J., Chen, X., Roberts, C. J., Mei, Y., Anderson, D. G., Langer, R., Alexander, M. R., Davies, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acrylate+polymers+with+hydro-responsive+topography.">Acrylate polymers with hydro-responsive topography.</a> Soft Matter. 7, 7194-9197 (2011).</li><li>Pernagallo, S., Unciti-Broceta, A., Diaz-Mochon, J. J., Bradley, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deciphering+cellular+morphology+and+biocompatibility+using+polymer+microarrays.">Deciphering cellular morphology and biocompatibility using polymer microarrays.</a> Biomedical Materials. 3, (2008).</li></ol>

We have nothing to disclose.

Polymer microarrays have been successfully used for the discovery of new materials by screening hundreds of novel polymer in a biological assay and identifying 'hit' materials that can subsequently be scaled up to useful devices. In this case, the surface characterization described may be employed subsequent to the biological assay and exclusively on the 'hit' materials to study those materials in more detail. This strategy may be of interest if HTSC is not available to the experimentalist employing this approach. However, to fully utilize polymer microarrays to study biological-material interactions the entire array of hundreds of materials should be analyzed prior to biological assays using HTSC methodologies, which can subsequently be used to observe general structure-function trends. 
Contact printing relies on the metal pin sliding up and down freely within the pin holder. Pin and pin holder cleanliness is paramount is ensuring printing occurs successfully and should be rigorously undertaken. Before commencing a printing run the appropriate movement of the pin within the pin holder can be tested by performing a dry run, with no monomers present. The cleaning step should continue until the pin movement is achieved reproducibly.
Considerable thought should go into the design of the monomer mixture. In order to easily produce a combinatorial library of polymers, hundreds of copolymers are formed by mixing a few monomers at different ratios. Typically we produce 576 member libraries as this forms a 24 x 24 array, which is suitable for the geometry of a glass slide. In order to produce a combinatorial library that explores the most combinatorial space the easiest method is to mix 24 monomers pairwise at a 2:1 ratio. Alternatively, the inclusion of compositional gradients within the array are useful for enabling the observations of trends, which allows optimal monomer compositions to be determined. As an example of this 22 monomers can be used as the first component in a co-monomer mixture that is sequentially diluted with 1 of 6 second components. If 5 dilutions are used, for example mixing the first and second components at ratios of 90:10, 75:25, 50:50, 25:75 and 10:90, this would result in 488 unique copolymer solutions. To bring the total up to 576, replicates of the homopolymers of the monomers used can be introduced, which often is an important reference sample. 576 monomer solutions should be dispensed into 2 384 well plates. For programming the robot it is easier to have two identical plates in terms of the position of the monomers, thus, the monomer solutions should be split evenly between the two plates.
A significant amount of time can be saved in the preparation of the source plates by the use of multichannel pipettes, and the design of source plates should be determined in order to exploit the use of the multichannel pipettes.
To achieve automated HTSC of the arrays the spot position must be successfully aligned with the characterization apparatus. Typically the pitch of an acrylate array is 500-1000 &mu;m and the polymer spot diameter is 300 &mu;m. Most X-Y stages have a resolution below 10 &mu;m, thus there is adequate tolerance for the surface characterization apparatus to reliably access the array positions once the correct dimensions have been input to the sample positioning software. The limitation to the automated positioning is in fact the accurate printing of the array. To ensure accurate printing it is important to prevent movement of the substrate on the printing stage either using vacuum suction or spring clamps together with appropriate slide dimensions (note that both a US and EU standard slide size exist). 
ToF-SIMS is an extremely surface sensitive technique that will observe any contamination on samples. Thus, upmost care must be taken to avoid contact with the surface. Samples should only be handled, but the surface of interest not contacted with, with clean gloves (preferably polyethylene) and with freshly cleaned tweezers. We typically wash with chloroform and hexane. Sample storage prior to measurements is best done in a sample holder that holds the slides apart, for example the 5 slide holder or 20 slide holder.
The arrays are designed specifically to be compatible with many biological assay formats and readouts, that is, the substrate used is a microscope slide ideally suited to fluorescence scanners and light microscopes. This means the format is well suited to exploring many material-biological interactions. Furthermore, the format allows hundreds of materials to be screened in parallel. This allows many more materials to be screened than conventional methods whereby each new material chemistry is screened individually. The increased scope for biological-material interactions allows for the elucidation of mechanisms of biological surface interactions, as well as finding the optimal material for a given application.

<table border="1">
	<tbody>
 	<tr>
 	<td>Name of the reagent/equipment</td>
 <td>Company</td>
 <td>Catalogue/Model number</td>
 </tr>
 <tr>
 	<td>Epoxy slides</td>
 <td>Genetix</td>
 <td>K2652</td>
 </tr>
 <tr>
 	<td>Contact printer</td>
 <td>Biodot</td>
 <td>XYZ3060 Platform</td>
 </tr>
 <tr>
 	<td>Metal pin</td>
 <td>ArrayIt</td>
 <td>946MP6B</td>
 </tr>
 <tr>
 	<td>ToF-SIMS instrument</td>
 <td>ION-TOF</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 	<td>XPS instrument</td>
 <td>Kratos</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 	<td>WCA apparatus</td>
 <td>Kr&uuml;ss </td>
 <td>DSA 100</td>
 </tr>
 <tr>
 	<td>AFM</td>
 <td>Bruker</td>
 <td>Dimension Icon</td>
 </tr>
 <tr>
 	<td>RPMI-1640 cell culture media</td>
 <td>Sigma-Aldrich</td>
 <td>R0883</td>
 </tr>
 <tr>
 	<td>SYTO17</td>
 <td>Invitrogen</td>
 <td>S-7579</td>
 </tr>
 </tbody>
</table>

polymer microarrays for high throughput discovery of biomaterials

The discovery of novel biomaterials that are optimized for a specific biological application is readily achieved using polymer microarrays, which allows a combinatorial library of materials to be screened in a parallel, high throughput format1. Herein is described the formation and characterization of a polymer microarray using an on-chip photopolymerization technique 2. This involves mixing monomers at varied ratios to produce a library of monomer solutions, transferring the solution to a glass slide format using a robotic printing device and curing with UV irradiation. This format is readily amenable to many biological assays, including stem cell attachment and proliferation, cell sorting and low bacterial adhesion, allowing the ready identification of 'hit' materials that fulfill a specific biological criterion3-5. Furthermore, the use of high throughput surface characterization (HTSC) allows the biological performance to be correlated with physio-chemical properties, hence elucidating the biological-material interaction6. HTSC makes use of water contact angle (WCA) measurements, atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). In particular, ToF-SIMS provides a chemically rich analysis of the sample that can be used to correlate the cell response with a molecular moiety. In some cases, the biological performance can be predicted from the ToF-SIMS spectra, demonstrating the chemical dependence of a biological-material interaction, and informing the development of hit materials5,3.

Watch this Scientific Journal Video about Polymer Microarrays for High Throughput Discovery of Biomaterials at JoVE.com

Polymer Microarrays for High Throughput Discovery of Biomaterials

A description of the formation of a polymer microarray using an on-chip photopolymerization technique. The high throughput surface characterization using atomic force microscopy, water contact angle measurements, X-ray photoelectron spectroscopy and time of flight secondary ion mass spectrometry and a cell attachment assay is also described.

The discovery of novel biomaterials that are optimized for a specific biological application is readily achieved using polymer microarrays, which allows a ...

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