ArticleMolecular biology of the cell2016
A novel method to accurately locate and count large numbers of steps by photobleaching.
Article in Molecular biology of the cell, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 27 papers.
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27 citing papers in PubMed.
- Bayesian analysis and efficient algorithms for single-molecule fluorescence data and step counting.Biophysical journal · 2025Article
- Foundation model for efficient biological discovery in single-molecule time traces.Nature methods · 2025Article
- Bayesian analysis and efficient algorithms for single-molecule fluorescence data and step counting.bioRxiv : the preprint server for biology · 2025Article
- FluoroTensor: Identification and tracking of colocalised molecules and their stoichiometries in multi-colour single molecule imaging via deep learning.Computational and structural biotechnology journal · 2024Article
- Single-molecule imaging reveals a direct role of CTCF's zinc fingers in SA interaction and cluster-dependent RNA recruitment.Nucleic acids research · 2024Article
- A Biophysics Toolbox for Reliable Data Acquisition and Processing in Integrated Force-Confocal Fluorescence Microscopy.ACS photonics · 2024Article
- Article
- MaxCal can infer models from coupled stochastic trajectories of gene expression and cell division.Biophysical journal · 2023Article
- Single-molecule counting applied to the study of GPCR oligomerization.Biophysical journal · 2022Review
- Modeling Non-additive Effects in Neighboring Chemically Identical Fluorophores.The journal of physical chemistry. B · 2022Article
- Gold Ion Beam Milled Gold Zero-Mode Waveguides.Nanomaterials (Basel, Switzerland) · 2022Article
- Diffraction-Limited Molecular Cluster Quantification with Bayesian Nonparametrics.Nature computational science · 2022Article
- Photobleaching step analysis for robust determination of protein complex stoichiometries.Molecular biology of the cell · 2021Article
- Estimating the dynamic range of quantitative single-molecule localization microscopy.Biophysical journal · 2021Article
- Article
- Dynamics of Ku and bacterial non-homologous end-joining characterized using single DNA molecule analysis.Nucleic acids research · 2021Article
- PySTACHIO: Python Single-molecule TrAcking stoiCHiometry Intensity and simulatiOn, a flexible, extensible, beginner-friendly and optimized program for analysis of single-molecule microscopy data.Computational and structural biotechnology journal · 2021Article
- Analyzing protein dynamics from fluorescence intensity traces using unsupervised deep learning network.Communications biology · 2020Article
- Live cell imaging of single RNA molecules with fluorogenic Mango II arrays.Nature communications · 2020Article
- Stoichiometric Analyses of Soluble CD4 to Native-like HIV-1 Envelope by Single-Molecule Fluorescence Spectroscopy.Cell reports · 2019Article
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5 authors.
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Abstract
Photobleaching event counting is a single-molecule fluorescence technique that is increasingly being used to determine the stoichiometry of protein and RNA complexes composed of many subunits in vivo as well as in vitro. By tagging protein or RNA subunits with fluorophores, activating them, and subsequently observing as the fluorophores photobleach, one obtains information on the number of subunits in a complex. The noise properties in a photobleaching time trace depend on the number of active fluorescent subunits. Thus, as fluorophores stochastically photobleach, noise properties of the time trace change stochastically, and these varying noise properties have created a challenge in identifying photobleaching steps in a time trace. Although photobleaching steps are often detected by eye, this method only works for high individual fluorophore emission signal-to-noise ratios and small numbers of fluorophores. With filtering methods or currently available algorithms, it is possible to reliably identify photobleaching steps for up to 20-30 fluorophores and signal-to-noise ratios down to ∼1. Here we present a new Bayesian method of counting steps in photobleaching time traces that takes into account stochastic noise variation in addition to complications such as overlapping photobleaching events that may arise from fluorophore interactions, as well as on-off blinking. Our method is capable of detecting ≥50 photobleaching steps even for signal-to-noise ratios as low as 0.1, can find up to ≥500 steps for more favorable noise profiles, and is computationally inexpensive.
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