ArticleBiophysical journal2019
Quantifying Protein Copy Number in Super Resolution Using an Imaging-Invariant Calibration.
Article in Biophysical journal, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 19 papers.
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Who cites it
19 citing papers in PubMed.
- Molecular mapping in DNA-PAINT via modified Gaussian Mixture Modeling.Nature communications · 2026Article
- Interplay between stochastic enzyme activity and microtubule stability drives detyrosination enrichment on microtubule subsets.Current biology : CB · 2023Article
- Quantitative Single-Molecule Localization Microscopy.Annual review of biophysics · 2023Review
- Myo19 tethers mitochondria to endoplasmic reticulum-associated actin to promote mitochondrial fission.Journal of cell science · 2023Article
- Quantification of Dark Protein Populations in Fluorescent Proteins by Two-Color Coincidence Detection and Nanophotonic Manipulation.The journal of physical chemistry. B · 2022Article
- Through the Eyes of Creators: Observing Artificial Molecular Motors.ACS nanoscience Au · 2022Review
- Small Peptide-Protein Interaction Pair for Genetically Encoded, Fixation Compatible Peptide-PAINT.Nano letters · 2021Article
- Quantitative Super-Resolution Microscopy to Assess Adhesion of Neuronal Cells on Single-Layer Graphene Substrates.Membranes · 2021Article
- A pairwise distance distribution correction (DDC) algorithm to eliminate blinking-caused artifacts in SMLM.Nature methods · 2021Article
- Diffusion of kinesin motors on cargo can enhance binding and run lengths during intracellular transport.Molecular biology of the cell · 2021Article
- The emergence of phase separation as an organizing principle in bacteria.Biophysical journal · 2021Review
- Challenges facing quantitative large-scale optical super-resolution, and some simple solutions.iScience · 2021Review
- A septin GTPase scaffold of dynein-dynactin motors triggers retrograde lysosome transport.The Journal of cell biology · 2021Article
- Activation and Regulation of Cytoplasmic Dynein.Trends in biochemical sciences · 2020Review
- The Generation of Dynein Networks by Multi-Layered Regulation and Their Implication in Cell Division.Frontiers in cell and developmental biology · 2020Review
- A Picture Worth a Thousand Molecules-Integrative Technologies for Mapping Subcellular Molecular Organization and Plasticity in Developing Circuits.Frontiers in synaptic neuroscience · 2020Article
- Axonal transport: Driving synaptic function.Science (New York, N.Y.) · 2019Review
- Nuclear pores as versatile reference standards for quantitative superresolution microscopy.Nature methods · 2019Article
- Single-molecule localization to study cytoskeletal structures, membrane complexes, and mechanosensors.Biophysical reviews · 2019Review
Corrections and comments
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Authors and funding
5 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The use of super-resolution microscopy in recent years has revealed that proteins often form small assemblies inside cells and are organized in nanoclusters. However, determining the copy number of proteins within these nanoclusters constitutes a major challenge because of unknown labeling stoichiometries and complex fluorophore photophysics. We previously developed a DNA-origami-based calibration approach to extract protein copy number from super-resolution images. However, the applicability of this approach is limited by the fact that the calibration is dependent on the specific labeling and imaging conditions used in each experiment. Hence, the calibration must be repeated for each experimental condition, which is a formidable task. Here, using cells stably expressing dynein intermediate chain fused to green fluorescent protein (HeLa IC74 cells) as a reference sample, we demonstrate that the DNA-origami-based calibration data we previously generated can be extended to super-resolution images taken under different experimental conditions, enabling the quantification of any green-fluorescent-protein-fused protein of interest. To do so, we first quantified the copy number of dynein motors within nanoclusters in the cytosol and along the microtubules. Interestingly, this quantification showed that dynein motors form assemblies consisting of more than one motor, especially along microtubules. This quantification enabled us to use the HeLa IC74 cells as a reference sample to calibrate and quantify protein copy number independently of labeling and imaging conditions, dramatically improving the versatility and applicability of our approach.
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