Application of Peak Intensity Analysis to Measurements of Protein Binding to Lipid Vesicles and Erythrocytes Using Fluorescence Correlation Spectroscopy: Dependence on Particle Size

Fluorescence correlation spectroscopy (FCS) is a sensitive analytical tool for investigation of processes accompanied by changes in the mobility of molecules and complexes. In the present work, peak intensity analysis (PIA) in combination with the solution stirring using FCS setup was applied to exp...

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Bibliographic Details
Published in:The Journal of membrane biology 2017-02, Vol.250 (1), p.77-87
Main Authors: Antonenko, Yuri N., Lapashina, Anna S., Kotova, Elena A., Ramonova, Alla A., Moisenovich, Mikhail M., Agapov, Igor I.
Format: Article
Language:English
Subjects:
Animals
Binding sites
Biochemistry
Biomedical and Life Sciences
Biotin
Cattle
Erythrocytes
Erythrocytes - metabolism
Human Physiology
Life Sciences
Lipids - chemistry
Liposomes - chemistry
Membranes
Particle Size
Protein Binding
Proteins
Proteins - chemistry
Proteins - metabolism
Ribosome Inactivating Proteins, Type 2 - chemistry
Ribosome Inactivating Proteins, Type 2 - metabolism
Ricin - chemistry
Ricin - metabolism
Spectrometry, Fluorescence
Spectrum analysis
Staining and Labeling
Toxins, Biological - chemistry
Toxins, Biological - metabolism
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Summary:Fluorescence correlation spectroscopy (FCS) is a sensitive analytical tool for investigation of processes accompanied by changes in the mobility of molecules and complexes. In the present work, peak intensity analysis (PIA) in combination with the solution stirring using FCS setup was applied to explore the interaction between fluorescently labeled protein ligands and corresponding receptors located on membranes. In the system composed of biotinylated liposomes and fluorescently labeled streptavidin as a ligand, PIA allowed us to determine the optimum receptor concentration and demonstrate the essential dependence of the binding efficacy on the length of the linker between the biotin group and the polar head group of the lipid. The binding was dependent on the size of liposomes which was varied by lipid extrusion through filters of different pore diameters. The sensitivity of the method was higher with the liposomes of larger sizes. The PIA approach can be applied not only to liposomes but also to relatively large objects, e.g., erythrocytes or Sepharose beads derivatized with lactose as a receptor for the binding of viscumin and ricin.
ISSN:0022-2631
1432-1424
DOI:10.1007/s00232-016-9938-6