Single-molecule measurements of transient biomolecular complexes through microfluidic dilution

Research output: Contribution to journalArticle

Abstract

Single-molecule confocal microscopy experiments require concentrations which are low enough to guarantee that, on average, less than one single molecule resides in the probe volume at any given time. Such concentrations are, however, significantly lower than the dissociation constants of many biological complexes which can therefore dissociate under single-molecule conditions. To address the challenge of observing weakly bound complexes in single-molecule experiments in solution, we have designed a microfluidic device that rapidly dilutes samples by up to one hundred thousand times, allowing the observation of unstable complexes before they dissociate. The device can interface with standard biochemistry laboratory experiments and generates a spatially uniform dilution that is stable over time allowing the quantification of the relative concentrations of different molecular species.

Details

Authors
  • Mathew H Horrocks
  • Luke Rajah
  • Peter Jönsson
  • Magnus Kjaergaard
  • Michele Vendruscolo
  • Tuomas P J Knowles
  • David Klenerman
External organisations
  • University of Cambridge
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Physical Chemistry

Keywords

  • Indicator Dilution Techniques, Microfluidic Analytical Techniques, Microscopy, Fluorescence, Oligonucleotides, Journal Article, Research Support, Non-U.S. Gov't
Original languageEnglish
Pages (from-to)6855-9
Number of pages5
JournalAnalytical Chemistry
Volume85
Issue number14
Publication statusPublished - 2013 Jul 16
Publication categoryResearch
Peer-reviewedYes
Externally publishedYes