Yu Zhongbo, Dulin David, Cnossen Jelmer, Köber Mariana, van Oene Maarten M, Ordu Orkide, Berghuis Bojk A, Hensgens Toivo, Lipfert Jan, Dekker Nynke H
Department of Bionanoscience, Kavli Institute of Nanoscience, Faculty of Applied Sciences, Delft University of Technology, Lorentzweg 1, 2628 CJ Delft, The Netherlands.
Rev Sci Instrum. 2014 Dec;85(12):123114. doi: 10.1063/1.4904148.
To study the behavior of biological macromolecules and enzymatic reactions under force, advances in single-molecule force spectroscopy have proven instrumental. Magnetic tweezers form one of the most powerful of these techniques, due to their overall simplicity, non-invasive character, potential for high throughput measurements, and large force range. Drawbacks of magnetic tweezers, however, are that accurate determination of the applied forces can be challenging for short biomolecules at high forces and very time-consuming for long tethers at low forces below ∼1 piconewton. Here, we address these drawbacks by presenting a calibration standard for magnetic tweezers consisting of measured forces for four magnet configurations. Each such configuration is calibrated for two commonly employed commercially available magnetic microspheres. We calculate forces in both time and spectral domains by analyzing bead fluctuations. The resulting calibration curves, validated through the use of different algorithms that yield close agreement in their determination of the applied forces, span a range from 100 piconewtons down to tens of femtonewtons. These generalized force calibrations will serve as a convenient resource for magnetic tweezers users and diminish variations between different experimental configurations or laboratories.
为了研究生物大分子在力作用下的行为以及酶促反应，单分子力谱技术的进展已被证明具有重要作用。磁镊是这些技术中最强大的技术之一，这得益于其整体的简单性、非侵入性、高通量测量的潜力以及较大的力范围。然而，磁镊的缺点在于，对于短生物分子在高力下准确测定施加的力具有挑战性，而对于低于约1皮牛顿的低力下的长系链则非常耗时。在此，我们通过提出一种磁镊校准标准来解决这些缺点，该校准标准由四种磁体配置的测量力组成。每种这样的配置针对两种常用的市售磁性微球进行校准。我们通过分析珠子的波动在时域和频域中计算力。通过使用不同算法验证所得的校准曲线，这些算法在确定施加的力方面产生了密切一致的结果，其范围从100皮牛顿到数十飞牛顿。这些通用的力校准将为磁镊用户提供便利资源，并减少不同实验配置或实验室之间的差异。
