Sevilla-Pym Angelica, Primrose William L, Luppi Bruno T, Bergmann Katrina, Hudson Zachary M
Department of Chemistry, The University of British Columbia, 2036 Main Mall, Vancouver, BC V6T 1Z1, Canada.
ACS Appl Mater Interfaces. 2024 Sep 4;16(35):46133-46144. doi: 10.1021/acsami.4c10311. Epub 2024 Aug 21.
Selective imaging of specific subcellular structures provides valuable information about the cellular microenvironment. Materials exhibiting thermally activated delayed fluorescence (TADF) are rapidly emerging as metal-free probes with long-lived emission for intracellular time-gated imaging applications. Polymers incorporating TADF emitters can self-assemble into luminescent nanoparticles, termed polymer dots (Pdots), and this strategy enables them to circumvent the limitations of commercial organelle trackers and small molecule TADF emitters. In this study, diblock copolymers comprised of a hydrophilic block containing organelle-targeting monomers and a hydrophobic TADF-active block were synthesized by ring-opening metathesis polymerization (ROMP). Oxanorbornene-based monomers incorporating morpholine and triphenylphosphonium groups for lysosome and mitochondria targeting, respectively, were also synthesized. ROMP by sequential addition yielded well-defined diblock copolymers with dispersities <1.28. To analyze the effect of tuning the hydrophilic corona on cellular viability and uptake, we prepared Pdots with poly(ethylene glycol) (PEG) and bis-guanidinium (BGN) coronas, resulting in limited and efficient cellular uptake, respectively. Red-emissive Pdots with BGN-based coronas and organelle-targeting functionality were obtained with quantum yields up to 12% in water under air. Colocalization analysis confirmed that lysosome and mitochondria labeling in live HeLa cells was accomplished within 2 h of incubation, affording Pearson's correlation coefficients of 0.37 and 0.70, respectively. The potential application of these Pdots for time-resolved imaging is highlighted by a proof of concept using time-gated spectroscopy, which effectively separates the delayed emission of the TADF Pdots from the background autofluorescence of biological serum.
对特定亚细胞结构进行选择性成像可提供有关细胞微环境的宝贵信息。具有热激活延迟荧光（TADF）的材料正迅速成为用于细胞内时间门控成像应用的无金属长寿命发射探针。包含TADF发射体的聚合物可自组装成发光纳米颗粒，称为聚合物点（Pdots），这种策略使它们能够规避商业细胞器追踪剂和小分子TADF发射体的局限性。在本研究中，通过开环易位聚合（ROMP）合成了由含有细胞器靶向单体的亲水性嵌段和疏水性TADF活性嵌段组成的二嵌段共聚物。还合成了分别含有吗啉和三苯基鏻基团用于靶向溶酶体和线粒体的基于氧化降冰片烯的单体。通过顺序添加进行的ROMP产生了分散度<1.28的明确二嵌段共聚物。为了分析调节亲水性冠层对细胞活力和摄取的影响，我们制备了具有聚乙二醇（PEG）冠层和双胍（BGN）冠层的Pdots，分别导致有限和高效的细胞摄取。在空气中，基于BGN冠层和具有细胞器靶向功能的红色发射Pdots在水中的量子产率高达12%。共定位分析证实，在活HeLa细胞中孵育2小时内即可完成溶酶体和线粒体标记，皮尔逊相关系数分别为0.37和0.70。使用时间门控光谱的概念验证突出了这些Pdots在时间分辨成像中的潜在应用，该技术有效地将TADF Pdots的延迟发射与生物血清的背景自发荧光分离。
