Single-molecule Force Studies
Wesley P. Wong, P.I.
Precise manipulation of single molecules is leading to remarkable insights in physics, chemistry, biology and medicine. However, widespread adoption of single-molecule techniques is impeded by equipment cost and the laborious nature of making measurements one molecule at a time. We are developing a new approach to solve these issues: massively parallel single-molecule force measurements using centrifugal force. This approach is realized in a novel instrument that we call the Centrifuge Force Microscope (CFM), in which objects in an orbiting sample are subjected to a calibration-free, macroscopically uniform force-field while their micro-to-nanoscopic motions are observed.
We have demonstrated high-throughput single-molecule force spectroscopy with this technique by performing thousands of rupture experiments in parallel, characterizing force-dependent unbinding kinetics of an antibody-antigen pair in minutes rather than days. Additionally, we have verified the force accuracy of the instrument by measuring the well-established DNA overstretching transition at 66 +/- 3 pN. With significant benefits in efficiency, cost, simplicity, and versatility, single-molecule centrifugation has the potential to expand single-molecule experimentation to a wider range of researchers and experimental systems.
Single-molecule centrifugation: a new approach for massively-parallel single-molecule manipulation
4:55 PM
relevent group publications
K. Halvorsen, W.P. Wong^, “Massively parallel single-molecule manipulation using centrifugal force,” Biophysical Journal – Letters, 98 (11), L53-L55 (2010). [pdf]
^corresponding author
We are developing a new approach for performing massively parallel single-molecule force measurements using centrifugal force. This is accomplished with a new instrument that we call the Centrifuge Force Microscope.
photo credit: John Chervinsky
Antibody-antigen unbinding kinetics obtained from massively-parallel single-molecule measurements made with the Centrifuge Force Microscope (CFM): Force-dependent unbinding of digoxigenin and its antibody. Force clamps ranging from hundreds of femtoNewtons to several picoNewtons were applied using the CFM (filled triangles), as well as with the optical trap (open triangles). Each CFM data point was obtained from a single experiment lasting a few minutes, while optical trap data was collected serially over a period of many hours. Histograms of the rupture times were fit with a decaying exponential to obtain the off-rate at each force (inset).