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Correlation of fluorescence microscopy, electron microscopy, and NanoSIMS stable isotope imaging on a single tissue section.
Communications Biology ( IF 5.2 ) Pub Date : 2020-07-09 , DOI: 10.1038/s42003-020-1095-x
Céline Loussert-Fonta 1 , Gaëlle Toullec 1 , Arun Aby Paraecattil 2 , Quentin Jeangros 3 , Thomas Krueger 1 , Stephane Escrig 1 , Anders Meibom 1, 4
Affiliation  

Correlative light and electron microscopy allows localization of specific molecules at the ultrastructural level in biological tissue but does not provide information about metabolic turnover or the distribution of labile molecules, such as micronutrients. We present a method to directly correlate (immuno)fluorescent microscopy, (immuno)TEM imaging and NanoSIMS isotopic mapping of the same tissue section, with nanometer-scale spatial precision. The process involves chemical fixation of the tissue, cryo sectioning, thawing, and air-drying under a thin film of polyvinyl alcohol. It permits to effectively retain labile compounds and strongly increases NanoSIMS sensitivity for 13C-enrichment. The method is illustrated here with correlated distribution maps of a carbonic anhydrase enzyme isotype, β-tubulin proteins, and 13C- and 15N-labeled labile micronutrients (and their anabolic derivates) within the tissue of a reef-building symbiotic coral. This broadly applicable workflow expands the wealth of information that can be obtained from multi-modal, sub-cellular observation of biological tissue.



中文翻译:


单个组织切片上荧光显微镜、电子显微镜和 NanoSIMS 稳定同位素成像的相关性。



相关光学和电子显微镜可以在生物组织的超微结构水平上定位特定分子,但不能提供有关代谢更新或不稳定分子(例如微量营养素)分布的信息。我们提出了一种以纳米级空间精度直接关联同一组织切片的(免疫)荧光显微镜、(免疫)TEM 成像和 NanoSIMS 同位素图谱的方法。该过程包括组织的化学固定、冷冻切片、解冻和在聚乙烯醇薄膜下的风干。它可以有效保留不稳定的化合物,并大大提高 NanoSIMS 对13 C 富集的灵敏度。这里通过碳酸酐酶同种型、β-微管蛋白以及13 C 和15 N 标记的不稳定微量营养素(及其合成代谢衍生物)在造礁共生珊瑚组织内的相关分布图来说明该方法。这种广泛适用的工作流程扩展了从生物组织的多模式、亚细胞观察中可以获得的丰富信息。

更新日期:2020-07-09
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