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首页> 外文期刊>Analytical chemistry >Microtubule-Targetable Fluorescent Probe: Site-Specific Detection and Super-Resolution Imaging of Ultratrace Tubulin in Microtubules of Living Cancer Cells
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Microtubule-Targetable Fluorescent Probe: Site-Specific Detection and Super-Resolution Imaging of Ultratrace Tubulin in Microtubules of Living Cancer Cells

机译：靶向微管的荧光探针：活癌细胞微管中超痕量微管蛋白的部位特异性检测和超分辨率成像

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摘要

Tubulins in microtubules have been recognized as potential targets in cancer chemotherapy for several years. However, their detection and imaging in living cells, especially following exposure to anticancer drugs, remains difficult to achieve. This difficulty is due to the very small cross section of microtubules and the very small changes in tubulin concentration involved. Photoswitchable fluorescent probes combined with the "super-resolution" fluorescence imaging technique present an exciting opportunity for site-specific detection and super-resolution imaging of specific microscopic populations, such as tubulin. In this study, a tubulin specific photoswitchable fluorescent probe (Tu-SP), that labels and detects ultratrace levels of tubulin in microtubules of living biosystems, was designed and evaluated. To realize super-resolution fluorescence imaging, the spiropyran derivative (SP), a classic photoswitch, was introduced to Tu-SP as a fluorophore. To detect ultratrace tubulin, Tu-SP employed the tubulin inhibitor, alkaloid colchicine (Tu), as a recognition unit. Tu-SP exhibited nearly nonintrinsic fluorescence before binding to tubulin, even if there were divalent metal ions and 375 nm lasers, respectively. After binding to tubulin, a dramatic increase in fluorescence was detected within milliseconds when irradiated at 375 nm, this increase is a result of the transformation of Tu-SP into a colored merocyanine (Tu-SP-1) with fluorescence. Tu-SP was successfully used for site-specific imaging of tubulin at a resolution of 20 +/- 5 nm in microtubules of living cancer cells. More importantly, the probe was suitable for site-specific and quantitative detection of trace tubulin in microtubules of living biological samples.
机译：微管中的微管蛋白已被认为是癌症化学疗法中的潜在靶标。然而，它们在活细胞中的检测和成像，尤其是在暴露于抗癌药物后，仍然难以实现。该困难是由于微管的横截面非常小，并且所涉及的微管蛋白浓度的变化很小。光开关荧光探针与“超分辨率”荧光成像技术的结合为特定微观人群（如微管蛋白）的位点特异性检测和超分辨率成像提供了令人兴奋的机会。在这项研究中，设计并评估了微管蛋白特异性光开关荧光探针（Tu-SP），该探针可标记和检测生物系统微管中微管蛋白的超痕量水平。为了实现超分辨率荧光成像，将经典的光电开关螺吡喃衍生物（SP）作为荧光团引入了Tu-SP。为了检测超微量微管蛋白，Tu-SP采用了微管蛋白抑制剂生物碱秋水仙碱（Tu）作为识别单元。即使分别存在二价金属离子和375 nm激光，Tu-SP在与微管蛋白结合之前也表现出几乎非内在的荧光。与微管蛋白结合后，当在375 nm处照射时，在毫秒内检测到荧光显着增加，这种增加是Tu-SP转变为带有荧光的有色花菁（Tu-SP-1）的结果。 Tu-SP已成功用于微管蛋白的位点特异性成像，分辨率为20 +/- 5 nm，位于活癌细胞的微管中。更重要的是，该探针适用于定点和定量检测活生物样品微管中的微量微管蛋白。

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