STEM-EELS tomography of biological materials

生物材料的 STEM-EELS 断层扫描

• 批准号: 577091-2022
• 负责人: Strauss, MikeM
• 金额: $ 3.28万
• 依托单位: McGill University
• 依托单位国家: 加拿大
• 项目类别: Alliance Grants
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=743437
• 关键词: STEM; EELS; tomography; biological; materials

Biological structures within a cell are difficult to assess at high resolution, which poses a critical limit to understanding a wide range of biological processes that are relevant to various human diseases. Light microscopy has made exceptional advancements in cell biology, enabling spatial and temporal resolution of biological molecules within single cells, however these techniques are limited in resolution and their ability to quantify protein density. Conventional electron microscopy can provide higher resolution of cell structures, but rely on the use of heavy metal stains to provide contrast. To circumvent these limitations, we plan to develop a 3D imaging technique to examine the elemental composition of unstained biological material in situ. We will develop and advance a technique known as scanning transmission electron microscopy in combination with electron tomography. This will allow us to obtain electron-energy loss spectroscopy (EELS) information of biological samples, that not only informs about the shape and structure of materials at exquisite resolution, but also reveals the chemical composition - allowing us to identify the nature of the molecules that we observe in addition to their shape and structure. This technique will combine the power and resolution of traditional electron microscopy with chemical information from the biological spectra, enabling detailed structural information without the use of conventional contrast enhancing stains. While common in material science research, several barriers have limited the use of STEM- EELS in revealing fine structural details of biological specimens, which we are now in a unique position to overcome to reveal unprecedented insight into the structure and composition of cells. We have previously demonstrated the feasibility of energy loss spectroscopy in resolving biological molecules using electron spectroscopic imaging by mapping phosphorus (abundant in nucleic acids) and nitrogen (present in all protein and nucleic acids) mapping. We will expand on this technique by collecting spectra at each pixel, and will obtain pixel specific spectra at multiple tilt angles to generate element-specific pixel resolution 3D information about our biological sample.

细胞内的生物结构很难以高分辨率进行评估，这对理解与各种人类疾病相关的广泛生物过程构成了关键限制。光学显微镜在细胞生物学方面取得了非凡的进步，实现了单细胞内生物分子的空间和时间分辨率，但这些技术在分辨率和量化蛋白质密度的能力方面受到限制。传统的电子显微镜可以提供更高分辨率的细胞结构，但依赖于使用重金属染色剂来提供对比度。为了规避这些限制，我们计划开发一种 3D 成像技术来原位检查未染色生物材料的元素组成。我们将开发和推进一种与电子断层扫描相结合的扫描透射电子显微镜技术。这将使我们能够获得生物样品的电子能量损失光谱（EELS）信息，不仅以精确的分辨率了解材料的形状和结构，而且还揭示了化学成分，使我们能够识别分子的性质除了它们的形状和结构之外，我们还观察到它们。该技术将传统电子显微镜的功率和分辨率与生物光谱的化学信息结合起来，无需使用传统的对比增强染色剂即可获得详细的结构信息。虽然在材料科学研究中很常见，但一些障碍限制了 STEM-EELS 在揭示生物样本的精细结构细节方面的应用，而我们现在处于一个独特的位置来克服这些障碍，以揭示对细胞结构和组成的前所未有的洞察。我们之前已经通过绘制磷（核酸中丰富的）和氮（存在于所有蛋白质和核酸中）图谱，证明了能量损失光谱在使用电子光谱成像解析生物分子方面的可行性。我们将通过收集每个像素的光谱来扩展这项技术，并将获得多个倾斜角度的像素特定光谱，以生成有关我们的生物样本的元素特定像素分辨率 3D 信息。