Massively Parallel Single-Molecule Manipulation Using Centrifugal Force
Massively Parallel Single-Molecule Manipulation Using Centrifugal Force
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DOI：
10.1016/j.bpj.2010.03.012
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发表时间：
2010-06-02
影响因子：
3.4
通讯作者：
Wong, Wesley P.
中科院分区：
文献类型：
--
作者：
Halvorsen, Ken;Wong, Wesley P.
对单分子的精确操纵已经在物理学、化学、生物学和医学方面带来了非凡的见解。然而，单分子技术的广泛采用受到设备成本和每次测量一个分子的费力性质的阻碍。我们已经解决了这些问题，通过开发一种方法，使大规模并行单分子力测量使用离心力。这种方法是在一种仪器中实现的，我们称之为离心力显微镜，在这种仪器中，轨道样本中的物体受到无校准的，宏观上均匀的力场，同时观察它们的微观到纳米级运动。我们展示了高通量的单分子力谱与这种技术进行了数以千计的破裂实验平行，表征力依赖的解结合动力学的抗体-抗原对在几分钟内，而不是几天。此外，我们通过在66 +/- 3 pN测量已确立的DNA过度拉伸转变来验证仪器的力准确度。单分子离心在效率、成本、简单性和多功能性方面具有显著优势，有可能将单分子实验扩展到更广泛的研究人员和实验系统。
Precise manipulation of single molecules has already led to remarkable insights in physics, chemistry, biology, and medicine. However, widespread adoption of single-molecule techniques has been impeded by equipment cost and the laborious nature of making measurements one molecule at a time. We have solved these issues by developing an approach that enables massively parallel single-molecule force measurements using centrifugal force. This approach is realized in an instrument that we call the centrifuge force microscope 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 demonstrate 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 verify 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.