Source 1review2016Analytical and Bioanalytical Chemistry
Toolkit/STORM
STORM
Also known as: stochastic optical reconstruction microscopy
Taxonomy: Technique Branch / Method. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
The supplied source summary states that the review explicitly covers SMLM, including STORM/dSTORM.
Usefulness & Problems
Why this is useful
STORM is presented as a super-resolution microscopy technique for single-molecule localization in image-based spatial proteomics contexts. The abstract groups it with methods achieving 10 to 20 nm spatial resolution.; super-resolution imaging; single-molecule localization with high spatial resolution; STORM is described in the supplied evidence as a super-resolution microscopy method used for microgel structural studies. It contributes to the review's goal of more complete nanoscale characterization.; super-resolution imaging of microgels; localization-based structural characterization; STORM is listed as a representative localization-microscopy approach discussed in the review.; super-resolution fluorescence imaging at the nanoscale
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STORM is presented as a super-resolution microscopy technique for single-molecule localization in image-based spatial proteomics contexts. The abstract groups it with methods achieving 10 to 20 nm spatial resolution.
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super-resolution imaging
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single-molecule localization with high spatial resolution
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STORM is described in the supplied evidence as a super-resolution microscopy method used for microgel structural studies. It contributes to the review's goal of more complete nanoscale characterization.
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super-resolution imaging of microgels
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localization-based structural characterization
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STORM is listed as a representative localization-microscopy approach discussed in the review.
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super-resolution fluorescence imaging at the nanoscale
Problem solved
It enables nanoscale spatial resolution for subcellular protein imaging.; improves spatial resolution for protein imaging at subcellular scale; It helps recover structural information about microgels at nanoscale resolution.; supports nanoscale structural readout of microgels; It is used to overcome diffraction-limited imaging by localizing single emitters for super-resolution reconstruction.; obtaining sub-diffraction spatial information from fluorescence microscopy
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It enables nanoscale spatial resolution for subcellular protein imaging.
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improves spatial resolution for protein imaging at subcellular scale
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It helps recover structural information about microgels at nanoscale resolution.
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supports nanoscale structural readout of microgels
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It is used to overcome diffraction-limited imaging by localizing single emitters for super-resolution reconstruction.
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obtaining sub-diffraction spatial information from fluorescence microscopy
Problem links
improves spatial resolution for protein imaging at subcellular scale
LiteratureIt enables nanoscale spatial resolution for subcellular protein imaging.
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It enables nanoscale spatial resolution for subcellular protein imaging.
obtaining sub-diffraction spatial information from fluorescence microscopy
LiteratureIt is used to overcome diffraction-limited imaging by localizing single emitters for super-resolution reconstruction.
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It is used to overcome diffraction-limited imaging by localizing single emitters for super-resolution reconstruction.
supports nanoscale structural readout of microgels
LiteratureIt helps recover structural information about microgels at nanoscale resolution.
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It helps recover structural information about microgels at nanoscale resolution.
Published Workflows
Objective: Develop and apply a targeted photoswitchable SNAP-tag substrate for specific labeling of cellular proteins and STORM nanoscopy of microtubules.
Why it works: The workflow couples a benzylguanine-targeted Cy3-Cy5 photoswitch to SNAP-tag fusion proteins, allowing specific covalent labeling of the structure of interest and then STORM reconstruction from single-emitter localization. The paper argues that the small size and stoichiometric labeling of SNAP-tag help avoid problems associated with antibody-based targeting.
benzylguanine-mediated targeting of a Cy3-Cy5 photoswitch to SNAP-tag fusion proteinssingle-emitter localization through repeated photoswitching for STORM reconstructionchemical probe synthesisSNAP-tag fusion protein labelingconfocal specificity checkSTORM imaging
Stages
- 1.Probe synthesis and in vitro SNAP reactivity check(library_build)
This stage establishes that the synthesized BG-Cy3-Cy5 probe can be produced and can react with SNAP-tag before cellular imaging experiments.
Selection: Obtain a photoswitchable benzylguanine probe that reacts with SNAP-tag
- 2.Cellular labeling specificity check(confirmatory_validation)
The authors use confocal colocalization with α-tubulin immunostaining to confirm that the chemical labeling marks the intended structure before super-resolution reconstruction.
Selection: Verify that BG-Cy3-Cy5 specifically labels β-tubulin-SNAP in fixed cells and localizes to microtubules
- 3.STORM imaging and reconstruction of labeled microtubules(confirmatory_validation)
This stage tests whether the targeted photoswitchable labeling strategy supports STORM reconstruction of microtubules below the diffraction limit.
Selection: Demonstrate nanoscale imaging performance on the labeled structure
Steps
- 1.Synthesize BG-Cy3-Cy5 from commercially available materialsengineered probe
Create a benzylguanine-linked Cy3-Cy5 photoswitchable substrate for SNAP-tag targeting
The probe must exist before its SNAP reactivity and cellular imaging utility can be tested.
- 2.Test in vitro reaction of BG-Cy3-Cy5 with SNAP-tagprobe and target tag
Verify that the synthesized probe reacts with SNAP-tag before cell-based experiments
In vitro reactivity is a lower-complexity prerequisite for interpreting later cellular labeling results.
- 3.Express β-tubulin-SNAP in U2OS cellscellular target construct
Place SNAP-tag on microtubules for targeted labeling
A SNAP-tag fusion target is required before fixed-cell labeling with BG-Cy3-Cy5 can be performed.
- 4.Fix cells and incubate with BG-Cy3-Cy5 followed by washinglabeling probe and target construct
Chemically label β-tubulin-SNAP in fixed cells
The paper emphasizes that post-fixation labeling is a prerequisite for compatibility with STORM.
- 5.Perform α-tubulin immunostaining control and confocal colocalization imaging
Confirm correct localization and specificity of SNAP-directed labeling before STORM
This control reduces the risk that later super-resolution structures reflect nonspecific labeling rather than microtubules.
- 6.Image labeled samples under STORM conditions and acquire 5000 frames
Collect photoswitching image data for single-molecule localization reconstruction
STORM acquisition follows confirmation of specific labeling so that the reconstruction can be interpreted as the target structure.
- 7.Reconstruct and filter single-molecule localization images
Generate super-resolved images and remove nonstructured background
Image reconstruction and filtering are required after acquisition to resolve nanoscale microtubule structure from the raw frame series.
Taxonomy & Function
Primary hierarchy
Technique Branch
Method: A concrete measurement method used to characterize an engineered system.
Mechanisms
single-molecule localizationstochastic emitter activation or detection for image reconstructionTarget processes
localizationImplementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationimplementation constraint: payload burdenoperating role: sensor
The abstract indicates that suitable fluorophores are required and that spectral overlap constrains practical implementation. It is discussed for live-cell multi-target imaging.; practical implementation is hampered by fluorophore limitations and spectral overlap; The payload only supports that STORM is a microscopy method; detailed setup requirements are not provided here.; requires super-resolution microscopy capability, with exact prerequisites not detailed in the payload
The abstract says slow image acquisition remains a barrier to high temporal resolution tracking of multiple protein targets in live cells.; slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; The supplied evidence does not support that STORM alone captures all relevant interaction, phase-behavior, or counterion-related properties.; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Source 2review2020Nature Communications
Source 3primary paper2023Preprints.org
Ranked Claims
Slow image acquisition in current super-resolution microscopy limits high temporal resolution tracking of multiple protein targets in live-cell imaging.
STED, PALM, and STORM can achieve 10 to 20 nm spatial resolution in single-molecule localization.
spatial resolution 10 to 20 nm
Scattering methods including small-angle neutron scattering are presented as core tools for microgel structural characterization.
Super-resolution microscopy methods including dSTORM and STORM are presented as relevant tools for resolving microgel network morphology and nanoscale structure.
In silico synthesis and modeling are relevant for connecting microgel network architecture to swelling and deswelling behavior.
The fuzzy-sphere model is used as a reference structural model for radial microgel morphology.
The review highlights nanobodies as a labeling strategy that reduces linkage error relative to conventional antibodies in super-resolution imaging.
The review groups PALM, STORM/dSTORM, and GSDIM under single-molecule localization microscopy.
The review discusses labeling chemistry, fluorophore photophysics, quantitative super-resolution, live-cell imaging, correlative microscopy, and analysis algorithms alongside core imaging modalities.
This review covers major super-resolution microscopy modality families including SIM, STED/RESOLFT, and single-molecule localization microscopy.
Approval Evidence
3 sources4 linked approval claimsfirst-pass slug storm
we have super-resolution microscopy techniques such as STED, PALM and STORM able to achieve 10 to 20 nm spatial resolution in single molecule localisation
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Explicitly supported in the supplied web research summary as a localization microscopy method discussed by the review for microgel structural studies.
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The supplied source summary states that the review explicitly covers SMLM, including STORM/dSTORM.
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limitation statementsupports
Slow image acquisition in current super-resolution microscopy limits high temporal resolution tracking of multiple protein targets in live-cell imaging.
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performance statementsupports
STED, PALM, and STORM can achieve 10 to 20 nm spatial resolution in single-molecule localization.
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method relevancesupports
Super-resolution microscopy methods including dSTORM and STORM are presented as relevant tools for resolving microgel network morphology and nanoscale structure.
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method family membershipsupports
The review groups PALM, STORM/dSTORM, and GSDIM under single-molecule localization microscopy.
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Comparisons
Source-stated alternatives
STED and PALM are named as alternative super-resolution methods.; The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
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STED and PALM are named as alternative super-resolution methods.
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The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Source-backed strengths
reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis
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reported as able to achieve 10 to 20 nm spatial resolution
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named by the review scaffold as part of the super-resolution characterization axis
Compared with 3D-dSTORM
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with direct stochastic optical reconstruction microscopy
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with dSTORM
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with GSDIM
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with PALM
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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STED and PALM are named as alternative super-resolution methods.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with photoactivated localization microscopy
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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STED and PALM are named as alternative super-resolution methods.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with photo-activation localization microscopy
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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STED and PALM are named as alternative super-resolution methods.
Source:
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with photoactivation localization microscopy
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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STED and PALM are named as alternative super-resolution methods.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with RESOLFT
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
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The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with reversible saturable optically linear transitions
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with small-angle neutron scattering
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
Compared with STED
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
STED and PALM are named as alternative super-resolution methods.
Source:
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with STED microscopy
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
STED and PALM are named as alternative super-resolution methods.
Source:
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with stimulated emission depletion microscopy
STED and PALM are named as alternative super-resolution methods.; The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
STED and PALM are named as alternative super-resolution methods.
Source:
The review also covers PALM, dSTORM, GSDIM, STED/RESOLFT, and SIM.
Compared with stochastic optical reconstruction microscopy
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
Shared frame: source-stated alternative in extracted literature
Strengths here: reported as able to achieve 10 to 20 nm spatial resolution; named by the review scaffold as part of the super-resolution characterization axis.
Relative tradeoffs: slow image acquisition limits high temporal resolution tracking of multiple protein targets in live cell imaging; the provided payload does not specify review-backed comparative strengths versus dSTORM or other methods.
Source:
The review scaffold contrasts or complements STORM with dSTORM, SANS, AFM, and modeling.
Ranked Citations
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