Super Resolution Imaging of Nanoparticles Cellular Uptake and Trafficking

Abstract

Understanding the interaction between synthetic nanostructures and living cells is of crucial importance for the development of nanotechnology-based intracellular delivery systems. Fluorescence microscopy is one of the most widespread tools owing to its ability to image multiple colors in native conditions. However, due to the limited resolution, it is unsuitable to address individual diffraction-limited objects. Here we introduce a combination of super-resolution microscopy and single-molecule data analysis to unveil the behavior of nanoparticles during their entry into mammalian cells. Two-color Stochastic Optical Reconstruction Microscopy (STORM) addresses the size and positioning of nanoparticles inside cells and probes their interaction with the cellular machineries at nanoscale resolution. Moreover, we develop image analysis tools to extract quantitative information about internalized particles from STORM images. To demonstrate the potential of our methodology, we extract previously inaccessible information by the direct visualization of the nanoparticle uptake mechanism and the intracellular tracking of nanoparticulate model antigens by dendritic cells. Finally, a direct comparison between STORM, confocal microscopy, and electron microscopy is presented, showing that STORM can provide novel and complementary information on nanoparticle cellular uptake.

Progress toward Understanding the Interactions between DNA Nanostructures and the Cell

“…We call for the delineation of the intracellular trafficking and exoyctosis of DNA at a higher spatial resolution afforded by conventional imaging tools, such as super‐resolution imaging. In 2016, van der Zwaag et al used stochastic optical construction microscopy to visualize individual nanoparticles inside cells and track single endocytic events …”

Cellular uptake of nanoparticles: journey inside the cell

“…10a illustrates lateral resolution improvements obtained from STORM imaging of polystyrene NPs internalized within HeLa cervical carcinoma cells. 207 Unlike confocal and conventional widefield methods, STORM can differentiate NPs of varying size (300 v 80 nm diameter) and functionality (color), providing previously unattainable information regarding aggregation state and location, as well as the capability to perform multiplex analyses in high-throughput assay formats.…”

“…i) Laser-ablation inductively coupled plasma mass spectrometry (ICP-MS) imaging (heatmap) of mouse fibroblasts (grayscale) incubated with Au (left) and Ag (right) nanoparticles. Reproduced with permission from (a) 207 , (b) 209 , (c) 243 , (d) 214 , (e) 217 , (f) 221 , (g) 225 , (h) 228 , and (i) 234 . Copyright (a) 2016 American Chemical Society, (b) 2014 Købler et al ., (c) 2015 Grant et al ., (d) 2014 Nature Publishing Group, (e) 2013 Nature Publishing Group, (f) 2014 Nature Publishing Group, (g) 2014 Nature Publishing Group, (h) 2016 Strohm et al ., and (i) 2012 American Chemical Society.…”

Analysing intracellular deformation of polymer capsules using structured illumination microscopy

“…Recently, stochastic optical reconstruction microscopy (STORM) and structured illumination microscopy (SIM) techniques have been applied for imaging the internalisation and intracellular processing of nanoparticles. [21][22][23] Herein we examine the intracellular deformation of polymer capsules in different cell lines using the super high resolution microscopy technique, SIM. SIM provides more than a two-fold increase in resolution by collecting information from the frequency space outside the observable region, and is able to clearly identify nanostructures (ca.…”