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An Automated Two-Dimensional Optical Force Clamp for Single Molecule Studies

Summary: An Automated Two-Dimensional Optical Force Clamp for Single
Molecule Studies
Matthew J. Lang,* Charles L. Asbury,* Joshua W. Shaevitz,
and Steven M. Block*
Departments of *Biological Sciences,
Physics, and
Applied Physics, Stanford University, Stanford, California 94305-5020 USA
ABSTRACT We constructed a next-generation optical trapping instrument to study the motility of single motor proteins,
such as kinesin moving along a microtubule. The instrument can be operated as a two-dimensional force clamp, applying
loads of fixed magnitude and direction to motor-coated microscopic beads moving in vitro. Flexibility and automation in
experimental design are achieved by computer control of both the trap position, via acousto-optic deflectors, and the sample
position, using a three-dimensional piezo stage. Each measurement is preceded by an initialization sequence, which includes
adjustment of bead height relative to the coverslip using a variant of optical force microscopy (to 4 nm), a two-dimensional
raster scan to calibrate position detector response, and adjustment of bead lateral position relative to the microtubule
substrate (to 3 nm). During motor-driven movement, both the trap and stage are moved dynamically to apply constant force
while keeping the trapped bead within the calibrated range of the detector. We present details of force clamp operation and
preliminary data showing kinesin motor movement subject to diagonal and forward loads.
Optical trapping has been used extensively to study the
motion of individual motor proteins such as kinesin, myo-


Source: Asbury, Chip - Department of Physiology and Biophysics, University of Washington at Seattle
Block, Steven - Departments of Biological Sciences & Applied Physics, Stanford University
Croquette, Vincent - Laboratoire de Physique Statistique, Département de Physique, École Normale Supérieure
Lang, Matthew - Division of Biological Engineering & Department of Mechanical Engineering, Massachusetts Institute of Technology (MIT)


Collections: Biology and Medicine; Biotechnology; Engineering; Physics