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Single-molecule binding experiments on long time scales

Journal Article


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Abstract


  • We describe an approach for performing single-molecule binding experiments on time scales from hours to days, allowing for the observation of slower kinetics than have been previously investigated by single-molecule techniques. Total internal reflection fluorescence microscopy is used to image the binding of labeled ligand to molecules specifically coupled to the surface of an optically transparent flow cell. Long-duration experiments are enabled by ensuring sufficient positional, chemical, thermal, and imagestability. Principal components of this experimental stability include illumination timing, solution replacement, and chemical treatment of solution to reduce photodamage and photobleaching; and autofocusing to correct for spatial drift.

Authors


Publication Date


  • 2010

Citation


  • Elenko, M. P., Szostak, J. W. & van Oijen, A. M. (2010). Single-molecule binding experiments on long time scales. Review of Scientific Instruments, 81 (8), 083705-1-083705-9.

Scopus Eid


  • 2-s2.0-77956317809

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=3214&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/2196

Has Global Citation Frequency


Start Page


  • 083705-1

End Page


  • 083705-9

Volume


  • 81

Issue


  • 8

Place Of Publication


  • United States

Abstract


  • We describe an approach for performing single-molecule binding experiments on time scales from hours to days, allowing for the observation of slower kinetics than have been previously investigated by single-molecule techniques. Total internal reflection fluorescence microscopy is used to image the binding of labeled ligand to molecules specifically coupled to the surface of an optically transparent flow cell. Long-duration experiments are enabled by ensuring sufficient positional, chemical, thermal, and imagestability. Principal components of this experimental stability include illumination timing, solution replacement, and chemical treatment of solution to reduce photodamage and photobleaching; and autofocusing to correct for spatial drift.

Authors


Publication Date


  • 2010

Citation


  • Elenko, M. P., Szostak, J. W. & van Oijen, A. M. (2010). Single-molecule binding experiments on long time scales. Review of Scientific Instruments, 81 (8), 083705-1-083705-9.

Scopus Eid


  • 2-s2.0-77956317809

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=3214&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/2196

Has Global Citation Frequency


Start Page


  • 083705-1

End Page


  • 083705-9

Volume


  • 81

Issue


  • 8

Place Of Publication


  • United States