Photonic-chip assisted correlative light and electron microscopy

Abstract

Correlative light and electron microscopy (CLEM) unifies the versatility of light microscopy (LM) with the high resolution of electron microscopy (EM), allowing one to zoom into the complex organization of cells. Here, we introduce photonic chip assisted CLEM, enabling multi-modal total internal reflection fluorescence (TIRF) microscopy over large field of view and high precision localization of the target area of interest within EM. The photonic chips are used as a substrate to hold, to illuminate and to provide landmarking of the sample through specially designed grid-like numbering systems. Using this approach, we demonstrate its applicability for tracking the area of interest, imaging the three-dimensional (3D) structural organization of nano-sized morphological features on liver sinusoidal endothelial cells such as fenestrations (trans-cytoplasmic nanopores), and correlating specific endo-lysosomal compartments with its cargo protein upon endocytosis.

Chip-based multimodal super-resolution microscopy for histological investigations of cryopreserved tissue sections

“…A simple thin layer of platinum deposited on top of the chip minimizes the charging effects and enables a good correlation between the light and the electron microscopy images. Importantly, the photonic chip can incorporate coordinate land-markings to facilitate the location and further correlation of the ROIs under study 36 . Lastly, the chip-based CLEM strategy presented here, in combination with the Tokuyasu method, can be executed within one working day from the sample sectioning steps to the SEM imaging, implying a significant time improvement as compared to the typical one-week imaging throughput associated with most CLEM approaches 41 .…”

“…The MMI illumination further aids in providing spatiotemporal sparsity that is useful for highly dense and heterogeneous samples such as tissues. Correlative imaging with other established methods including electron microscopy 36 and quantitative phase microscopy 27 can be seamlessly implemented on the photonic chip, expanding the opportunities both for routine analysis and for basic histology research. The photonic chip-based microscopy can be implemented on standard optical microscopy platforms upon a few adaptations for the integration of a photonic chip module (Fig.…”

“…Correlative imaging with other established methods including electron microscopy 36 and quantitative phase microscopy 27 can be seamlessly implemented on the photonic chip, expanding the opportunities both for routine analysis and for basic histology research.…”

Extracellular vesicles through the blood–brain barrier: a review

“…This will allow the colocalization of two signals, one in the lipophilic space, such as with an organic fluorescent dye, and another bioluminescence enzyme system or a nanoparticle marker in the hydrophilic space, assuring the EV structure integrity once the visualization experiment is performed. Another possibility for characterization in this context could be Correlative Light and Electron Microscopy (CLEM) or Cryo-CLEM [ 82 , 83 ]. This will allow a dynamic description of the motion of EVs from the periphery, through the BBB, and to specific anatomical structures of the CNS, although the precise mechanisms of transcytosis would have to be evaluated as previously described.…”

UV photonic integrated circuits for far-field structured illumination autofluorescence microscopy

“…PICs have already sparked a tremendous number of applications of paramount importance in telecommunications 11 , healthcare 12 , biological research 13 , quantum information and computation 14 , 15 , metrology 16 , 17 , chemical 18 , 19 and biological sensing 20 , environmental monitoring 21 , and artificial intelligence 22 . With regard to super-resolution optical microscopy, PICs have been mainly used for near-field imaging, in particular for implementing wide field-of-view waveguide-based total internal reflection fluorescence (TIRF) microscopy combined with direct stochastic reconstruction techniques 23 , 24 , fluorescence-fluctuation-based techniques 25 , and wide field-of-view waveguide-based points accumulation in nanoscale topography (PAINT) 26 . The operating wavelength range of current PICs extends from visible to mid-infrared, but does not yet includes the UV range extending from 200 to 400 nm.…”