Photothermal confocal multicolor microscopy of nanoparticles and nanodrugs in live cells

Growing biomedical applications of non-fluorescent nanoparticles (NPs) for molecular imaging, disease diagnosis, drug delivery, and theranostics require new tools for real-time detection of nanomaterials, drug nano-carriers, and NP-drug conjugates (nanodrugs) in complex biological environments without additional labeling. Photothermal (PT) microscopy (PTM) has enormous potential for absorption-based identification and quantification of non-fluorescent molecules and NPs at a single molecule and 1.4 nm gold NP level. Recently, we have developed confocal PTM providing three-dimensional (3D) mapping and spectral identification of multiple chromophores and fluorophores in live cells. Here, we summarize recent advances in the application of confocal multicolor PTM for 3D visualization of single and clustered NPs, alone and in individual cells. In particular, we demonstrate identification of functionalized magnetic and gold–silver NPs, as well as graphene and carbon nanotubes in cancer cells and among blood cells. The potential to use PTM for super-resolution imaging (down to 50 nm), real-time NP tracking, guidance of PT nanotherapy, and multiplex cancer markers targeting, as well as analysis of non-linear PT phenomena and amplification of nanodrug efficacy through NP clustering and nano-bubble formation are also discussed.

无荧光纳米颗粒（non-fluorescent nanoparticles, NPs）在分子成像、疾病诊断、药物递送及治疗诊断学领域的生物医学应用日益增多，亟需开发新型工具，以在复杂生物环境中对纳米材料、药物纳米载体以及纳米药物偶联物（nanodrugs）实现无需额外标记的实时检测。光热显微镜（Photothermal microscopy, PTM）在基于吸收原理的单分子及1.4纳米金纳米颗粒级别无荧光分子与纳米颗粒的鉴定与定量方面具备巨大应用潜力。近期，我们研发了共聚焦光热显微镜，可实现活细胞内多种生色团与荧光团的三维（3D）成像及光谱鉴定。本文总结了多色共聚焦光热显微镜在单个及聚集态纳米颗粒（无论单独存在还是位于单个细胞内）的三维可视化方面的最新应用进展。具体而言，我们验证了功能化磁性、金-银纳米颗粒以及石墨烯与碳纳米管在癌细胞及血细胞中的鉴定效果。此外，本文还探讨了光热显微镜的多项应用潜力：包括分辨率低至50纳米的超分辨成像、纳米颗粒的实时追踪、光热纳米治疗的导航、多重癌症靶点标记、非线性光热现象的分析，以及通过纳米颗粒聚集与纳米气泡形成增强纳米药物疗效的相关研究。