Laboratoire de Bioimagerie et Pathologies, UMR 7021, CNRS, University of Strasbourg, FR 67401 Illkirch, France.
Université de Paris, Institute of Psychiatry and Neuroscience of Paris (IPNP), INSERM U1266, "Membrane Traffic in Healthy and Diseased Brain", F 75014 Paris, France.
Bioconjug Chem. 2020 Mar 18;31(3):875-883. doi: 10.1021/acs.bioconjchem.0c00023. Epub 2020 Feb 26.
The plasma membrane (PM) plays a major role in many biological processes; therefore, its proper fluorescence staining is required in bioimaging. Among the commercially available PM probes, styryl dye FM1-43 is one of the most widely used. In this work, we demonstrated that fine chemical modifications of FM1-43 can dramatically improve the PM staining. The newly developed probes, SP-468 and SQ-535, were found to display enhanced photophysical properties (reduced cross-talk, higher brightness, improved photostability) and, unlike FM1-43, provided excellent and immediate PM staining in 5 different mammalian cell types including neurons (primary culture and tissue imaging). Taking advantage of these features, we successfully used SP-468 in STED super resolution neuronal imaging. Additionally, we showed that the new probes displayed differences in their internalization pathways compared to their parent FM1-43. Finally, we showed that the new probes kept the ability to stain the PM of plant cells. Overall, this work presents new useful probes for PM imaging in cells and tissues and provides insights on the molecular design of new PM targeting molecules.
质膜（PM）在许多生物过程中起着重要作用；因此，在生物成像中需要对其进行适当的荧光染色。在市售的 PM 探针中，荧光素 FM1-43 是应用最广泛的探针之一。在这项工作中，我们证明了 FM1-43 的精细化学修饰可以显著改善 PM 染色。新开发的探针 SP-468 和 SQ-535 表现出增强的光物理性质（减少串扰、更高的亮度、提高光稳定性），与 FM1-43 不同，在包括神经元在内的 5 种不同的哺乳动物细胞类型（原代培养和组织成像）中能够立即提供出色的 PM 染色。利用这些特性，我们成功地在 STED 超分辨率神经元成像中使用了 SP-468。此外，我们还表明，与母体 FM1-43 相比，这些新探针在其内化途径上存在差异。最后，我们证明了这些新探针能够保持对植物细胞 PM 的染色能力。总的来说，这项工作为细胞和组织中的 PM 成像提供了新的有用探针，并为新的 PM 靶向分子的分子设计提供了新的见解。
