Format

Send to

Choose Destination
Methods. 2010 Oct;52(2):192-200. doi: 10.1016/j.ymeth.2010.06.010. Epub 2010 Jun 8.

Single-molecule FRET of protein-nucleic acid and protein-protein complexes: surface passivation and immobilization.

Author information

1
Department of Chemistry, Wayne State University, Detroit, MI 48202, USA.

Abstract

Single-molecule fluorescence spectroscopy reveals the real time dynamics that occur during biomolecular interactions that would otherwise be hidden by the ensemble average. It also removes the requirement to synchronize reactions, thus providing a very intuitive approach to study kinetics of biological systems. Surface immobilization is commonly used to increase observation times to the minute time scale, but it can be detrimental if the sample interacts non-specifically with the surface. Here, we review detailed protocols to prevent such interactions by passivating the surface or by trapping the molecules inside surface immobilized lipid vesicles. Finally, we discuss recent examples where these methods were applied to study the dynamics of important cellular processes at the single-molecule level.

PMID:
20554047
PMCID:
PMC3321382
DOI:
10.1016/j.ymeth.2010.06.010
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center