Short summary for those who wish to use SAXS:

The resolution of SAXS is up to about 1.5 nm, the method can be used for macromolecules with MW from about 5 kDa up to 100 MDa.

One can study low resolution shape, quaternary structure, complex formation, add missing portions to high resolution models, analyze oligomeric mixtures and kinetic processes, and responses to changes conditions like pH, ionic strength, temperature etc.

For structure analysis (shape, quaternary structure), monodisperse samples are required. Monodispersity must be checked before doing SAXS by DLS, native gel, ultracentrifugation etc.

About 2-3 mg of purified material is needed for a complete study. The typical concentration range starts from about 0.5-1 mg/ml to about 10 mg/ml. At X33, the sample volume per measurement is currently about 60-70µl (manual filling) or 90-100µl (automated sample changer). We strongly recommend to use the automated mode.

FPLC available for X33: Users have the possibility to use an on-site FPLC system prior to measurement (AKTA purifier, GE life sciences). A gel-filtration run on samples is advised for difficult samples to separate aggregates and higher oligomers from the protein to be studied. It is also possible to use this system to analyse fractions from gel-filtration and characterise oligomeric states of individual proteins or the formation/dissociation of complexes.

For the supplied Superose-6 10/300 column, the recommended loading concentration is ~ 10 mg/ml and a loading volume of 200 - 500 µl. One should then obtain fractions in the 1 mg/ml concentration range and a sample volume of 50-100 µl used for SAXS. A typical run, including column equilibration is approximately 2 hours. Users are encouraged to bring 1 litre of buffer for column equilibration and the subsequent run, however, when this is not possible buffers can be prepared on site.

If this service is required users must request beam-time during the week. No support for the wet-lab is available on weekends.

Beamline automation: the sample changer presently allows for unattended measurements. Remote control is currently being tested and will be offered to the users in test mode as of summer 2009. For remote/Fedex operation, all samples must be fully prepared, without the necessity of dilutions or additions. More detailed instructions will be posted in due course.

If you are interested in a collaborative proposal, EMBL staff will be happy to help you not only with data collection and processing, but also with the interpretation and model building.

Apply for the SAXS beam time.

Learn more about data collection and processing.