Non-invasive light scattering methods provide information about the size and the homogeneity of particle populations in solution. The mean translational diffusion coefficient, the hydrodynamic radius, and the polydispersity of a particle population can be determined in dynamic mode. The mean particle mass and the second virial coefficient characterizing the interactions between particles can be determined in static mode. Applications include the detection of aggregates and the titration of biological macromolecules with ligands.
Contact: Philippe BENAS
Dynamic light scattering (DLS) analyses record the fluctuations of the intensity of light scattered due to Brownian motion. A correlator generates an autocorrelation function which is then deconvoluted in an intensity particle size distribution. A translational diffusion coefficient (Dt) can be extracted when the sample is a single population of identical particles. The standard deviation on Dt informs about sample polydispersity. A hydrodynamic radius can be derived from Dt assuming that particles are hard spheres. Extrapolation to zero concentration (D0) eliminates the contribution of interactions (see e.g. Lorber et al. 2012, Biochem Mol Biol Educ 40, 372-82). Measurements can be done at any temperatures between 5°C and 50°C with appropriate cells.
In static light scattering (SLS) the intensity of the scattered light is a function of particle mass and concentration. The mean mass of the particles in a homogeneous sample can be derived from measurements done at various concentrations.
How it works
The sample is contained in a cuvette. It can be recovered for further analyses.
- Large aggregates which scatter light much more than smaller particles should be removed either by size-exclusion column chromatography or by ultracentrifugation. Filtration is not recommended for biological macromolecules.
- Sample volumes and concentrations vary with instrument sensitivity and particle size.
- In DLS, the diffusion coefficient should be corrected for the absolute (dynamic) viscosity and the refractive index of the solvent. In SLS, the increment of particle refractive index and the precise particle concentration should be known.
Additional equipment is available to determine properties of the solvent. It includes:
- an Anton Paar AMVn micro-viscosimeter for measuring the kinematic viscosity,
- a 5 mL glass pycnometer for the determination of the density, and
- an Abbe refractometer for measuring refractive indices.
Send enquiries about appointments and prices to Bernard Lorber.
ITC (Isothermal Titration Calorimetry) is the gold standard for measuring biomolecular interactions. ITC simultaneously determines all binding parameters (n, K, ΔH and ΔS) in a single experiment – information that cannot be obtained from any other method.
Contact: Eric Ennifar et Karl Brillet
The switchSENSE technology mainly allows to measure the interaction between biomolecules thanks to the association and dissociation kinetics.
Other important biological parameters can be explored such as hydrodynamic radius, conformational changes, enzymatic activity…
The technology is based on the immobilization of a ligand at the end of a double strand of nucleic acid that oscillates under the effect of an alternating electric current.
Coupled with a microfluidic technique, this approach allows to work with small amounts of biological material (50 pmoles on average) and a very wide range of molecular weight (a few kDa to several MDa).
Sequencing apparatus allowing the detection of fluorescent cDNAs
- Equipment: ABI 3130xI Genetic Analyser (Applied Biosystems)
- This device allows a high throuput detection for:
– The mapping of RNA molecules in order to study their structure
– The footprinting analysis of proteins or other molecule on RNA
Principle of the technique:
– RNA is modified by specific enzymatic digestion or chemical modification (SHAPE reagents, DMS, EDC, …)
– Modifications are detected as stops during reverse transcription of a 5’-fluorescently labeled primer.
VIC (560 nm)
NED (580 nm)
For information on how to prepare your samples or if you plan to perform a new analysis contact Valérie Vivet-Boudou.
Contact: Valérie VIVET-BOUDOU
French press type disintegrator for large cell volumes.
Laminar flow hoods, CO2 ovens, phase contrast microscope.
Contact: Jean-Christophe Paillart
Microbiology laboratory of containment class 2, allowing the manipulation of microorganisms belonging to group 2. Example: Staphyloccocus aureus, Shigella sp.
Compulsory training before handling in L2 provided by Anne-Catherine HELFER-LE FOLL.
Equipement : PSM type II (= microbiological safety station including an air suction system that keeps the work surface permanently depressed), incubator, centrifuge, secure entry by code door.
Contact: Anne-Catherine HELFER-LE FOLL
PowerPoint of the training: « How to manipulate in L2 lab »