ArticleAnalytical biochemistry2013
Are fluorescence-detected sedimentation velocity data reliable?
Article in Analytical biochemistry, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 15 papers.
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Who cites it
15 citing papers in PubMed, 23 citations in OpenAlex.
- Long-Term Stable Biosensing Using Multiscale Biostructure-Preserving Metal Thin Films.Biosensors · 2026Article
- Sedimentation velocity FDS studies of antibodies in pooled human serum.European biophysics journal : EBJ · 2023Article
- Analysis of nonideality: insights from high concentration simulations of sedimentation velocity data.European biophysics journal : EBJ · 2020Article
- AUC measurements of diffusion coefficients of monoclonal antibodies in the presence of human serum proteins.European biophysics journal : EBJ · 2018Article
- Use of fluorescence-detected sedimentation velocity to study high-affinity protein interactions.Nature protocols · 2017Article
- Sedimentation of Reversibly Interacting Macromolecules with Changes in Fluorescence Quantum Yield.Biophysical journal · 2017Article
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- MCM ring hexamerization is a prerequisite for DNA-binding.Nucleic acids research · 2015Article
- Combining biophysical methods for the analysis of protein complex stoichiometry and affinity in SEDPHAT.Acta crystallographica. Section D, Biological crystallography · 2015Article
- Accounting for photophysical processes and specific signal intensity changes in fluorescence-detected sedimentation velocity.Analytical chemistry · 2014Article
- Analysis of protein interactions with picomolar binding affinity by fluorescence-detected sedimentation velocity.Analytical chemistry · 2014Article
- Effect of basic cell-penetrating peptides on the structural, thermodynamic, and hydrodynamic properties of a novel drug delivery vector, ELP[V5G3A2-150].Biochemistry · 2014Article
- Tools for the quantitative analysis of sedimentation boundaries detected by fluorescence optical analytical ultracentrifugation.PloS one · 2013Article
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Corrections and comments
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Authors and funding
5 authors at 3 institutions in 1 country.
Funding
Abstract
Sedimentation velocity analytical ultracentrifugation is a classical biophysical technique that is commonly used to analyze the size, shape, and interactions of biological macromolecules in solution. Fluorescence detection provides enhanced sensitivity and selectivity relative to the standard absorption and refractrometric detectors, but data acquisition is more complex and can be subject to interference from several photophysical effects. Here, we describe methods to configure sedimentation velocity measurements using fluorescence detection and evaluate the performance of the fluorescence optical system. The fluorescence detector output is linear over a concentration range of at least 1 to 500nM fluorescein and Alexa Fluor 488. At high concentrations, deviations from linearity can be attributed to the inner filter effect. A duplex DNA labeled with Alexa Fluor 488 was used as a standard to compare sedimentation coefficients obtained using fluorescence and absorbance detectors. Within error, the sedimentation coefficients agree. Thus, the fluorescence detector is capable of providing precise and accurate sedimentation velocity results that are consistent with measurements performed using conventional absorption optics, provided the data are collected at appropriate sample concentrations and the optics are configured correctly.
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