Mazloom-Farsibaf Hanieh, Farzam Farzin, Fazel Mohamadreza, Wester Michael J, Meddens Marjolein B M, Lidke Keith A
Department of Physics and Astronomy, University of New Mexico, Albuquerque, New Mexico, United States of America.
Department of Mathematics and Statistics, University of New Mexico, Albuquerque, New Mexico, United States of America.
PLoS One. 2021 Jan 28;16(1):e0246138. doi: 10.1371/journal.pone.0246138. eCollection 2021.
Visualizing actin filaments in fixed cells is of great interest for a variety of topics in cell biology such as cell division, cell movement, and cell signaling. We investigated the possibility of replacing phalloidin, the standard reagent for super-resolution imaging of F-actin in fixed cells, with the actin binding peptide 'lifeact'. We compared the labels for use in single molecule based super-resolution microscopy, where AlexaFluor 647 labeled phalloidin was used in a dSTORM modality and Atto 655 labeled lifeact was used in a single molecule imaging, reversible binding modality. We found that imaging with lifeact had a comparable resolution in reconstructed images and provided several advantages over phalloidin including lower costs, the ability to image multiple regions of interest on a coverslip without degradation, simplified sequential super-resolution imaging, and more continuous labeling of thin filaments.
在固定细胞中可视化肌动蛋白丝对于细胞生物学中的各种主题，如细胞分裂、细胞运动和细胞信号传导，具有极大的研究价值。我们研究了用肌动蛋白结合肽“lifeact”替代鬼笔环肽（固定细胞中F-肌动蛋白超分辨率成像的标准试剂）的可能性。我们比较了用于基于单分子的超分辨率显微镜的标记物，其中AlexaFluor 647标记的鬼笔环肽用于dSTORM模式，Atto 655标记的lifeact用于单分子成像、可逆结合模式。我们发现，使用lifeact成像在重建图像中具有相当的分辨率，并且相对于鬼笔环肽具有多个优势，包括成本更低、能够在不降解的情况下对盖玻片上的多个感兴趣区域进行成像、简化的连续超分辨率成像以及对细丝进行更连续的标记。
