Microscopy Research and Technique virtual issue: "Correlative light and electron microscopy".

Abstract

Correlative Light and Electron Microscopy (CLEM) has become increasingly important for the analysis of the structure, function, and dynamics of cells as well as for surface analysis in material science. This combination of multiple imaging technologies provides more data than each stand-alone technique. CLEM intended for Biological Science includes a growing collection of methods that allows the localization of a molecule of interest within a high-resolution cellular map but, as we show here, is not limited to standard fluorescence and TEM anymore. This special issue of Microscopy Research and Technique (MRT) contains 11 articles selected from Biological and Material Science papers published from 1998 to 2023 that represent innovative applications of correlative light and electron microscopy across a wide spectrum of scientific research. These articles were of great interest to us and we hope that they will be to you as well. There is a focus on the use of probes in a majority of these papers so we will accompany this virtual issue with a review on CLEM probes. 1. Braet, F., Wisse, E., Bomans, P., Frederik, P., Geerts, W., Koster, A., Soon, L., & Ringer, S. (2007). Contribution of high-resolution correlative imaging techniques in the study of the liver sieve in three-dimensions. Microscopy Research and Technique, 70(3), 230–242. https://doi.org/10.1002/jemt.20408. In this paper, the authors compare different correlative high-resolution imaging methodologies and how these microscopy techniques facilitate the accumulation of new insights in the morpho-functional and structural organization of the hepatic sieve showing the involvement of special domains in the de novo formation and disappearance of hepatic fenestrae. 2. Hand, G. M., Martone, M. E., Stelljes, A., Ellisman, M. H., & Sosinsky, G. E. (2001). Specific labeling of connexin43 in NRK cells using tyramide-based signal amplification and fluorescence photooxidation. Microscopy Research and Technique, 52(3), 331–343. https://doi.org/10.1002/1097-0029(20010201)52:3<331::AID-JEMT1017>3.0.CO;2-H. In this paper, the authors use correlative microscopy to localize connexin pools or structures in gap junctions and discovered that other cellular substructures interact with gap junction proteins. The use of tyramide signal amplification techniques and fluorescence photooxidation allowed for three-dimensional localization of proteins using high-voltage electron microscopy. 3. Hein, L. R. D. O., de Oliveira, J. A., & de Campos, K. A. (2013). Correlative light-electron fractography for fatigue striations characterization in metallic alloys. Microscopy Research and Technique, 76(9), 909–913. https://doi.org/10.1002/jemt.22247. In this paper, the authors describe a correlative light-electron fractography technique that combined correlative microscopy to the extended depth-from-focus reconstruction method for the analysis of fatigue striations spacing. The correlative fractography technique demonstrated a large potential for quantitative analysis of fracture surfaces and provided a comprehensive approach in failure analysis. 4. Nishiyama, H., Teramoto, K., Suga, M., & Sato, C. (2014). Positively charged nanogold label allows the observation of fine cell filopodia and flagella in solution by atmospheric scanning electron microscopy. Microscopy Research and Technique, 77(2), 153–160. https://doi.org/10.1002/jemt.22322. In this paper, the authors describe a method in which positively charged nanogold particles were used to label the surfaces of bacteria and cultured animal cells, exploiting their net negative surface charge. After gold enhancement to increase the size of the nanogold particles, atmospheric scanning electron microscopy (ASEM) imaging of the bacteria in aqueous solution revealed pili and delicate spiral flagella, in contrast with images of dried flagella recorded by standard SEM. 5. Powell, R. D., Halsey, C. M., & Hainfeld, J. F. (1998). Combined fluorescent and gold immunoprobes: reagents and methods for correlative light and electron microscopy. Microscopy Research and Technique, 42(1), 2–12. https://doi.org/10.1002/(SICI)1097-0029(19980701)42:1<2::AID-JEMT2>3.0.CO;2-Y. In this paper, the authors describe novel immunoprobes that incorporated a fluorescent label and a 1.4 nm gold cluster to label cellular structures. The fluorescent and gold cluster probes enabled a new degree of correlation between fluorescence and electron microscopy and could also be used to check labeling of specimens before processing for electron microscopy. 6. Robinson, J. M., Takizawa, T., Vandré, D. D., & Burry, R. W. (1998). Ultrasmall immunogold particles: Important probes for immunocytochemistry. Microscopy Research and Technique, 42(1), 13–23. https://doi.org/10.1002/(SICI)1097-0029(19980701)42:1<13::AID-JEMT3>3.0.CO;2-S. In this paper, the authors review the immunocytochemical literature and compare conventional colloidal gold and ultrasmall gold particles as immunoprobes. They also discuss the relative advantages and disadvantages of each of these types of particles for immunocytochemical applications and provide examples of the use of a fluorescently labeled ultrasmall immunoprobe for correlative microscopy. 7. Rodighiero, S., Torre, B., Sogne, E., Ruffilli, R., Cagnoli, C., Francolini, M., Di Fabrizio, E., & Falqui, A. (2015). Correlative scanning electron and confocal microscopy imaging of labeled cells coated by indium-tin-oxide. Microscopy Research and Technique, 78(6), 433–443. https://doi.org/10.1002/jemt.22492. In this paper, the authors describe a method that combined confocal microscopy imaging of cells membranes with scanning electron microscopy. They demonstrated that a thin layer of indium-tin-oxide (ITO) could be effectively deposited by ion sputtering on cells previously tagged with both fluorophores and gold particles targeting the same surface molecules. The addition of ITO to SEM samples provided a stable electrical conductivity and preservation of the back scattered electron signal coming from the gold-conjugated markers on the cell surface. 8. Sherman, S., Nachmias, D., & Elia, N. (2015). A simple, straightforward correlative live-cell-imaging-structured-illumination-microscopy approach for studying organelle dynamics. Microscopy Research and Technique, 78(9), 777–783. https://doi.org/10.1002/jemt.22540. In this paper, the authors presented an easily applicable assay for studying mitochondrial membrane fission in cells. Using a correlative approach that combined structured illumination Microscopy (SIM) with spinning disk (SD) microscopy, they imaged the cells at physiological conditions with most suitable imaging recording protocol, without compromising the quality of the high-resolution 3D SIM data. 9. Vicidomini, G., Gagliani, M.C., Cortese, K., Krieger, J., Buescher, P., Bianchini, P., Boccacci, P., Tacchetti, C., & Diaspro, A. (2010). A novel approach for correlative light electron microscopy analysis. Microscopy Research and Technique, 73(3), 215–224. https://doi.org/10.1002/jemt.20777. In this paper, the authors developed a new CLEM method optimized for cryosections (Tokuyasu method) based on a robust specimen preparation protocol, and on an innovative image processing toolbox for a novel type of multimodal analysis. This method has been validated on correlative analysis of Russel Bodies compartments and opened new possibilities for high-throughput correlative light electron microscopy. 10. Quintana, C., Wu, T. D., Delatour, B., Dhenain, M., Guerquin-Kern, J. L., & Croisy, A. (2007). Morphological and chemical studies of pathological human and mice brain at the subcellular level: correlation between light, electron, and nanosims microscopies. Microscopy Research and Technique, 70(4), 281–295. https://doi.org/10.1002/jemt.20403. In this paper, the authors reported the use of the NanoSIMS-50™ ion microprobe that allowed the simultaneous identification of five elements with high sensitivity, in combination with transmission electron microscopy in Alzheimer brains. Using this correlative technique, they demonstrated the subcellular distribution of iron–ferritin–hemosiderin in the hippocampus of Alzheimer disease patients, and the presence of Ca–Fe mineralized amyloid deposits in the thalamus of transgenic mice. 11. Badar, F., & Xia, Y. (2022). The interface region between articular cartilage and bone by μMRI and PLM at microscopic resolutions. Microscopy Research and Technique, 85, 1483–1493. https://analyticalsciencejournals.onlinelibrary.wiley.com/doi/epdf/10.1002/jemt.24011. The last paper in this list is also the most recent. The authors use microscopic MRI and polarized light microscopy to perform multimodal imaging of cartilage and bone. The paper nicely demonstrates the breadth of correlative microscopy techniques and the integration of new microscopy techniques in the workflows. Katia Cortese: Conceptualization; writing – original draft; writing – review and editing. Paul Verkade: Conceptualization; writing – original draft; writing – review and editing. Data sharing not applicable to this article as no datasets were generated or analysed during the current study.