Fluorescence Correlation Spectroscopy

National facility

The FCS facility guides users in measurements of concentrations and sizes of biomolecules as well as interactions between biomolecules, in solution or in living cells. We offer access to the latest developments in FCS and related techniques to all researchers at universities in Sweden. Support is given in the design and planning of experiments as well as in performing measurements and interpreting and analyzing the experimental data. Projects are usually carried out as scientific collaborations between the respective research groups and the FCS facility. Two of the techniques that can be utilized are powerful variants of FCS that recently were developed by the facility staff: Inverse FCS and FRET-FCS. Inverse FCS determines the absolute size of receptor oligomers or domains, even on highly dynamic membranes which is not possibly with any current technique. FRET-FCS enables high sensitivity to detect small subpopulations of FRET-active oligomers.

SERVICES

  • Analysis and interpretation of experimental data.
  • Project planning and experimental design of FCS-measurements.
  • Support in specimen preparation.
  • Training in FCS-techniques.

APPLICATIONS

  • Dynamic analysis of receptor-receptor or ligand-receptor interactions on living cell membranes.
  • Analysis of protein-protein interactions in the cytoplasm or nucleus of living cells.
  • Analysis of protein-carrying vesicles in solution, to determine whether different proteins are coupled to one and the same vesicle.
  • protein-peptide interactions and affinity estimations at nM concentrations in solution.
  • Aggregation analysis in solution at nM concentrations.

 

 

EQUIPMENT

  • Classic FCS.
  • Dual colour cross-correlation (FCCS).
  • FRET-FCS. Allows sensitive detection of FRET-active oligomers present at small fractions.
  • Image correlation spectroscopy (ICS).
  • Inverse FCS. Determines the absolute volume of nanoparticles in solution, or the size of clusters/domains in cell membranes.
  • SICS. A label-free technique which in combination with FCS allows simultaneous analysis of labeled and unlabeled nanoparticles in solution.

RECENT PROJECTS

Services/collaborative projects:

  • Agonist-induced dimer dissociation as a macromolecular step in G protein-coupled receptor signaling, Petersen, J., ..., Schulte, G., Nature Communications, 2017, vol. 8, article number 226.
  • Spontaneous Vesiculation and pH-Induced Disassembly of a Lysosomotropic Detergent: Impacts on Lysosomotropism and Lysosomal Delivery, 2016, Langmuir, 32, 13566-13575.
  • Membrane raft integrity constrains basal insulin secretion, but is impaired in type 2-diabetes. Nagaraj, V., ..., Renström, E., Molecular Endocrinology2016, 30, 1059-1069.
  • A novel labelling approach for nano cellulose fibers. Navarro et al, Biomacromolecules, 201617, 1101-1109.
  • Detection of rare Amyloid Beta oligomers in solution using FRET-FCS, Anal. Chem., 2015, 87 (23), pp 11700–11705
  • Analysis of pH-driven dimerization of the N-terminal domain of spider silk proteins in solution. Kronqvist, N. et al, Nature Communications, 2014, vol. 5, article number 3254.
  • Analysis of nanoparticles used for purification of sewage plant water – detection of size-differences before and after water treatment. Lakshmanan, R. et al, 2014, Langmuir, 30, 1036−1044.

Technology development:

  • Scanning Inverse Fluorescence Correlation Spectroscopy (siFCS), Bergstrand et al, Opt. Express, 2014, 22 (11), 13073-13090.
  • FRET-FCS for detection of rare FRET-active oligomers, Wennmalm et al, Anal. Chem., 2015, 87 (23), 11700-11705 (Technology development within a collaborative project).
  • Label-Free Fluctuation Spectroscopy Based on Coherent Anti-Stokes Raman Scattering from Bulk Water Molecules, Rabasovic et al, ChemPhysChem, 2016, 17, 1025-1033.
  • Label-free analysis of nanoparticles based on scattering and interference, Wennmalm and Widengren, 2012J. Am. Chem. Soc., 134, 19516−19519.