Previous Article |
Table of Contents
| Next Article 
BIOLOGICAL SCIENCES / BIOPHYSICS
Selective aluminum passivation for targeted immobilization of single DNA polymerase molecules in zero-mode waveguide nanostructures
Pacific Biosciences, 1505 Adams Drive, Menlo Park, CA 94025
Communicated by Watt W. Webb, Cornell University, Ithaca, NY, November 20, 2007 (received for review August 6, 2007)
Optical nanostructures have enabled the creation of subdiffraction detection volumes for single-molecule fluorescence microscopy. Their applicability is extended by the ability to place molecules in the confined observation volume without interfering with their biological function. Here, we demonstrate that processive DNA synthesis thousands of bases in length was carried out by individual DNA polymerase molecules immobilized in the observation volumes of zero-mode waveguides (ZMWs) in high-density arrays. Selective immobilization of polymerase to the fused silica floor of the ZMW was achieved by passivation of the metal cladding surface using polyphosphonate chemistry, producing enzyme density contrasts of glass over aluminum in excess of 400:1. Yields of single-molecule occupancies of 
30% were obtained for a range of ZMW diameters (70–100 nm). Results presented here support the application of immobilized single DNA polymerases in ZMW arrays for long-read-length DNA sequencing.
fluorescence | metal passivation | microscopy | polyvinyl phosphonic acid | single molecule
Conflict of interest statement: The authors are employed by Pacific Biosciences, Inc., which has carried out the research described as part of a for-profit commercial development program.
This article contains supporting information online at www.pnas.org/cgi/content/full/0710982105/DC1.
*To whom correspondence should be addressed. E-mail sturner{at}pacificbiosciences.com
© 2008 by The National Academy of Sciences of the USA
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg What's this?
