Artifact Formation in Single Molecule Localization Microscopy

Jm, Reichel; Vomhof, Thomas; Michaelis, Jens

doi:10.1101/700955

Cited by 2 publications

(2 citation statements)

References 46 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…3 , 4 , and S7 ). These slight differences are likely attributable to a small amount of noise after correction ( 43 ).…”

Section: Discussionmentioning

confidence: 99%

“…Hoogendoorn et al. previously implemented a more complex temporal median filter using Python ( 29 ) that has been applied in STORM analysis tools such as FIRESTORM ( 43 ) and the commercial software, Huygens Localizer (Scientific Volume Imaging, Hilversum, the Netherlands). This technique is more complicated relative to the use of the simple method tested here, and although it performs similarly with simulated data, its ability to correct for macrophage autofluorescence is less accurate.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Corrected Super-Resolution Microscopy Enables Nanoscale Imaging of Autofluorescent Lung Macrophages

Ambrose

Dechantsreiter

Montero

et al. 2020

Biophysical Journal

View full text Add to dashboard Cite

Observing the cell surface and underlying cytoskeleton at nanoscale resolution using super-resolution microscopy has enabled many insights into cell signaling and function. However, the nanoscale dynamics of tissue-specific immune cells have been relatively little studied. Tissue macrophages, for example, are highly autofluorescent, severely limiting the utility of light microscopy. Here, we report a correction technique to remove autofluorescent noise from stochastic optical reconstruction microscopy (STORM) data sets. Simulations and analysis of experimental data identified a moving median filter as an accurate and robust correction technique, which is widely applicable across challenging biological samples. Here, we used this method to visualize lung macrophages activated through Fc receptors by antibody-coated glass slides. Accurate, nanoscale quantification of macrophage morphology revealed that activation induced the formation of cellular protrusions tipped with MHC class I protein. These data are consistent with a role for lung macrophage protrusions in antigen presentation. Moreover, the tetraspanin protein CD81, known to mark extracellular vesicles, appeared in ring-shaped structures (mean diameter 93 5 50 nm) at the surface of activated lung macrophages. Thus, a moving median filter correction technique allowed us to quantitatively analyze extracellular secretions and membrane structure in tissue-derived immune cells.

show abstract

“…3 , 4 , and S7 ). These slight differences are likely attributable to a small amount of noise after correction ( 43 ).…”

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Corrected Super-Resolution Microscopy Enables Nanoscale Imaging of Autofluorescent Lung Macrophages

Ambrose

Dechantsreiter

Montero

et al. 2020

Biophysical Journal

View full text Add to dashboard Cite

show abstract

Analysis of super-resolution single molecule localization microscopy data: A tutorial

Fazel

Wester

2022

AIP Advances

View full text Add to dashboard Cite

The diffraction of light imposes a fundamental limit on the resolution of light microscopes. This limit can be circumvented by creating and exploiting independent behaviors of the sample at length scales below the diffraction limit. In super-resolution single molecule localization microscopy (SMLM), the independence arises from individual fluorescent labels stochastically switching between dark and fluorescent states, which in turn allows the pinpointing of fluorophores post experimentally using a sequence of acquired sparse image frames. Finally, the resulting list of fluorophore coordinates is utilized to produce high resolution images or to gain quantitative insight into the underlying biological structures. Therefore, image processing and post-processing are essential stages of SMLM. Here, we review the latest progress on SMLM data processing and post-processing.

show abstract

Artifact Formation in Single Molecule Localization Microscopy

Cited by 2 publications

References 46 publications

Corrected Super-Resolution Microscopy Enables Nanoscale Imaging of Autofluorescent Lung Macrophages

Corrected Super-Resolution Microscopy Enables Nanoscale Imaging of Autofluorescent Lung Macrophages

Analysis of super-resolution single molecule localization microscopy data: A tutorial

Contact Info

Product

Resources

About