SEC-MALS couples a size exclusion chromatography and different detectors, static light scattering, refractive index, absorbance and Dynamic light scattering or viscometer. It combines the separation of macromolecules and their characterization.
Application
- Determination of concentration, molecular weight and hydrodynamic radius
- Determination of aggregation and oligomerisation states: homogeneity of the sample
- Determination of conformational changes: interactions
- Determination of the stoichiometry of complexes, including for complex systems associating protein-detergent, protein-RNA, protein-polymer… It is a method often appropriated for studying interactions of membrane protein and/or glycosylated proteins.
Staff/ Contact
Manager : Caroline Mas
Operated by : Aline Le Roy and Caroline Mas
Scientific expert: Christine EBEL
Office: IBS room 355 / CIBB room 017
PAOL: IBS room 327
OMNISEC: CIBB room 001
Specific equipment
Protein Analysis On Line: PAOL (Shimadzu & Wyatt)
- HPLC (Shimadzu)
- Detector: UV (PDA) (Shimadzu), MALS (miniDAWN TREOS, Wyatt), DLS (Dynapro Nanostar, Wyatt), RI (Optilab rEX, Wyatt)
- Auto sampler (Shimadzu): 4 – 40°C
- Collector (Shimadzu): 4 – 20°
- Column oven (Winsep): 4 – 30°C
- Used software: LC Solution (Shimadzu); ASTRA and Dynamics (Wyatt).
OMNISEC (Malvern)
- Detectors: RALS, LALS, UV (PDA), RI, Viscometer
- Auto sampler: 4 – 40°C
- Column oven: 25 – 35 °C
- Detector oven: 25-35°C
- Used software: OMNISEC (Malvern)
- Silica column: KW 802.5, KW 803 and KW 804 (Shodex) and WTC050N5 (Wyatt)
- Agarose/Dextran column: Superdex 30, Superdex 75, Superdex 200 and Superose 6 10/300 GL (GE Healthcare)
How to make a request?
These instruments work in service mode.
Each new project manager should contact us.
Users can be trained to the analysis if the research project requires extensive use of the instrument.
Users agree to comply with the general terms of Biophysics and SEC-MALS condition of use.
All applications should be accompanied by a request form. The applicant should fill in the first part, which is completed in agreement by both part, in order to define the best way to conduct the experiments, before signature of the parts.
Samples
The sample is typically at 1-10 mg/mL; the typical volume is 50 µl. The samples must be centrifuge during 5-10 min 16 000g, or filtered at 0.1µm. Samples are not stored –unless specific request.
The elution buffer must be filtered at 0.1µm. It contains typically 100mM salt; silica columns require a pH in the range 3 – 7.5. Our standard buffer is PBS: 30mM Na Phosphate pH 6.8, 100 mM NaCl, 0.3 g/L NaN3.
