ArticleNature communications2022
Absolute protein quantification using fluorescence measurements with FPCountR.
Article in Nature communications, 2022. 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.
- Stabilization of microbial cell population for long-term plasmid production in continuous biomanufacturing.Bioprocess and biosystems engineering · 2026Article
- Assessment and detection of nucleotide excision repair capability in human cells: a critical review of current functional methods.Molecular biology reports · 2026Review
- BindPred: a framework for predicting protein-protein binding affinity from language model embeddings.Bioinformatics (Oxford, England) · 2026Article
- Absolute quantification of fluorescent protein fusions by mass spectrometry.Protein science : a publication of the Protein Society · 2026Article
- Noise-Guided Design of Synthetic Protein Waves in Living Cells.ACS synthetic biology · 2026Article
- Making GFP count: a validated framework for absolute protein quantification in precision fermentation.Applied microbiology and biotechnology · 2026Article
- Dynamic Robust Control of Microbial Communities Using Cybergenetics.Methods in molecular biology (Clifton, N.J.) · 2026Article
- Characterization of Synthetic Gene Circuits with Absolute Quantification in Continuous Culture.Methods in molecular biology (Clifton, N.J.) · 2026Article
- Article
- Synthetic gene circuits in plants: recent advances and challenges.Quantitative plant biology · 2025Review
- In Vivo Quantification of Surfactin Nonribosomal Peptide Synthetase Complexes inMicroorganisms · 2024Article
- Mutational profiling of SARS-CoV-2 papain-like protease reveals requirements for function, structure, and drug escape.Nature communications · 2024Article
- Novel delivery systems for controlled release of bacterial therapeutics.Trends in biotechnology · 2024Review
- Quantitative measurement of cell-surface displayed proteins based on split-GFP assembly.Microbial cell factories · 2024Article
- Data hazards in synthetic biology.Synthetic biology (Oxford, England) · 2024Review
- Standardization of Fluorescent Reporter Assays in Synthetic Biology across the Visible Light Spectrum.ACS synthetic biology · 2023Article
- Parsley: a web app for parsing data from plate readers.Bioinformatics (Oxford, England) · 2023Article
- Structural Insights into the Binding of Red Fluorescent Protein mCherry-Specific Nanobodies.International journal of molecular sciences · 2023Article
- Advancing reproducibility can ease the 'hard truths' of synthetic biology.Synthetic biology (Oxford, England) · 2023Review
Corrections and comments
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Authors and funding
2 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
This paper presents a generalisable method for the calibration of fluorescence readings on microplate readers, in order to convert arbitrary fluorescence units into absolute units. FPCountR relies on the generation of bespoke fluorescent protein (FP) calibrants, assays to determine protein concentration and activity, and a corresponding analytical workflow. We systematically characterise the assay protocols for accuracy, sensitivity and simplicity, and describe an 'ECmax' assay that outperforms the others and even enables accurate calibration without requiring the purification of FPs. To obtain cellular protein concentrations, we consider methods for the conversion of optical density to either cell counts or alternatively to cell volumes, as well as examining how cells can interfere with protein counting via fluorescence quenching, which we quantify and correct for the first time. Calibration across different instruments, disparate filter sets and mismatched gains is demonstrated to yield equivalent results. It also reveals that mCherry absorption at 600 nm does not confound cell density measurements unless expressed to over 100,000 proteins per cell. FPCountR is presented as pair of open access tools (protocol and R package) to enable the community to use this method, and ultimately to facilitate the quantitative characterisation of synthetic microbial circuits.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.