Instituto de Biofísica, Universidade Federal do Rio de Janeiro/Polo Xerém, Duque de Caxias 25245-390, Brazil.
J Mol Recognit. 2012 May;25(5):256-61. doi: 10.1002/jmr.2189.
The atomic force microscopy (AFM) has been used as a force sensor to measure unbinding forces of single bound complexes in the nanonewton and piconewton range. Force spectroscopy measurements can be applied to study both intermolecular and intramolecular interactions of complex biological and synthetic macromolecules. Although the AFM has been extensively used as a nano force sensor, the commercially available cantilever is limited to silicon and silicon nitride. Those materials reduce the adhesion sensitivity with specific surface and/or molecule. Here, we functionalized the AFM tip with carboxylic groups by applying acrylic acid (AA) vapor at radio frequency plasma treatment at 100 W for 5 min. This method provides a remarkable sensitivity enhancement on the functional group interaction specificity. The functionalized tip was characterized by scanning electron microscopy. The electron beam high resolution images have not shown significant tip sharpness modification. Silicon wafers (1 0 0)-no treated and functionalized by AA plasma treatment-were characterized by Auger electron spectroscopy to elucidate the silicon surface sputtering and demonstrate functionalization. The Fourier transform-infrared spectroscopy spectrum shows a high absorbance of avidin protein over the silicon surface functionalized by AA plasma treatment.We carried out force spectroscopy assay to measure the unbinding force between the well-established pair biotin-avidin. At pulling speed of 2 µm/s, we measured the unbinding force of 106 ± 23 pN, which is in good agreement with the literature, demonstrating the effectiveness of the tip functionalization by AA plasma treatment in biological studies.
原子力显微镜（AFM）已被用作力传感器，用于测量纳牛顿和皮牛顿范围内的单结合复合物的解结合力。力谱测量可用于研究复杂生物和合成大分子的分子间和分子内相互作用。尽管 AFM 已被广泛用作纳米力传感器，但市售的悬臂梁仅限于硅和氮化硅。这些材料降低了与特定表面和/或分子的附着力灵敏度。在这里，我们通过在 100 W 的射频等离子体处理下施加丙烯酸（AA）蒸气，将 AFM 尖端功能化为羧基。这种方法显著提高了官能团相互作用的特异性灵敏度。通过扫描电子显微镜对功能化尖端进行了表征。电子束高分辨率图像并未显示尖端锐度有明显改变。硅片（100）-未经处理和 AA 等离子体处理功能化-通过俄歇电子能谱进行了表征，以阐明硅表面溅射并证明功能化。傅里叶变换-红外光谱显示，经 AA 等离子体处理功能化的硅表面上吸附蛋白的吸收率很高。我们进行了力谱测定，以测量生物素-亲和素这一对成熟配对物之间的解结合力。在 2 µm/s 的拉动速度下，我们测量到的解结合力为 106 ± 23 pN，与文献值吻合良好，证明了 AA 等离子体处理对尖端功能化在生物研究中的有效性。
