Practical axial optical trapping

Rev Sci Instrum. 2012 Oct;83(10):103106. doi: 10.1063/1.4757862.

Abstract

We describe a new method for calibrating optical trapping measurements in which tension is applied in the direction of the laser beam to a molecule tethered between a surface and an optically trapped bead. Specifically, we present a generally-applicable procedure for converting from the measured scattering intensity and the measured stage displacement to applied tension and bead-coverslip separation, using measurements of the light intensity scattered from an untethered, trapped bead. Our calibration accounts for a number of effects, including aberrations and the interference of forward-reflected bead-scattered light with the trapping beam. To demonstrate the accuracy of our method, we show measurements of the DNA force-versus-extension relation using a range of laser intensities, and show that these measurements match the expected extensible wormlike-chain (WLC) behavior. Finally, we also demonstrate a force-clamp, in which the tension in a tether is held fixed while the extension varies as a result of molecular events.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Biomechanical Phenomena
  • Calibration
  • DNA / chemistry
  • DNA / metabolism
  • Lasers
  • Optical Tweezers*
  • Scattering, Radiation
  • Signal Processing, Computer-Assisted

Substances

  • DNA