Department of Chemical Engineering, McGill University, 3610 University Street, Montreal, Quebec H3A 0C5, Canada.
Environ Sci Technol. 2022 May 17;56(10):6426-6435. doi: 10.1021/acs.est.1c08480. Epub 2022 Apr 26.
Understanding of nanoplastic prevalence and toxicology is limited by imaging challenges resulting from their small size. Fluorescence microscopy is widely applied to track and identify microplastics in laboratory studies and environmental samples. However, conventional fluorescence microscopy, due to diffraction, lacks the resolution to precisely localize nanoplastics in tissues, distinguish them from free dye, or quantify them in environmental samples. To address these limitations, we developed techniques to label nanoplastics for imaging with stimulated emission depletion (STED) microscopy to achieve resolution at an order of magnitude superior to conventional fluorescence microscopy. These techniques include (1) passive sorption; (2) swell incorporation; and (3) covalent coupling of STED-compatible fluorescence dyes to nanoplastics. We demonstrate that our labeling techniques, combined with STED microscopy, can be used to resolve nanoplastics of different shapes and compositions as small as 50 nm. The longevity of dye labeling is demonstrated in different media and conditions of biological and environmental relevance. We also test STED imaging of nanoplastics in exposure experiments with the model worm . Our work shows the value of the method for detection and localization of nanoplastics as small as 50 nm in a whole animal without disruption of the tissue. These techniques will allow more precise localization and quantification of nanoplastics in complex matrices such as biological tissues in exposure studies.
对纳米塑料流行情况和毒理学的了解受到其尺寸小而导致的成像挑战的限制。荧光显微镜广泛应用于实验室研究和环境样本中追踪和识别微塑料。然而，由于衍射，传统荧光显微镜缺乏在组织中精确定位纳米塑料、将其与游离染料区分开来或在环境样本中对其进行定量的分辨率。为了解决这些限制，我们开发了用于具有受激发射损耗（STED）显微镜成像的纳米塑料标记技术，以实现比传统荧光显微镜优越一个数量级的分辨率。这些技术包括（1）被动吸附；（2）溶胀结合；和（3）与 STED 兼容的荧光染料与纳米塑料的共价偶联。我们证明，我们的标记技术与 STED 显微镜结合使用，可以分辨出小至 50nm 的不同形状和组成的纳米塑料。在不同的生物和环境相关的介质和条件下，证明了染料标记的耐久性。我们还在模式蠕虫的暴露实验中测试了纳米塑料的 STED 成像。我们的工作表明，该方法在不破坏组织的情况下，在整个动物中检测和定位小至 50nm 的纳米塑料具有价值。这些技术将允许在暴露研究中对生物组织等复杂基质中的纳米塑料进行更精确的定位和定量。
