Introduction

1) Getting beamtime (proposals, reports and deadlines): Access to beamtime at ESRF and ILL is granted via electronic proposal systems ( links see above). Potential users requesting beamtime must submit a scientific proposal online, providing scientific background on the experiments, samples to be measured, time requested etc. For more information, see ( links). The systems are generally open twice a year with deadlines that must be respected. In a proposal, a local contact that accompanies you during the experiment must be indicated in general. If successful, beamtime is generally attributed during the 6 months that follow the proposal deadline. The submitted proposals are reviewed by scientific committees that evaluate their scientific quality. Due to an overload factor that varies from instrument to instrument, not all proposals can be accepted.

2) Before the experiment (sample preparation and pre-characterization): The key to a successful SAXS/SANS experiment in biology is to know the biochemistry of the system studied. As a matter of fact, prior to a SAXS/SANS experiment, as much information as possible should be collected on the system investigated by complementary methods (gel filtration, light scattering, analytical ultracentrifugation, optical density, etc…). The following points are particularly important:

- Aggregation state: structural studies on aggregated samples are in general not possible, it is therefore essential to avoid aggregation. SANS practitioners should be aware that an elevated D2O content in the solution (as required for some experiments, link to contrast matching) may favor aggregation. In general, higher sample concentrations also favor aggregation. In such cases it is advisable to run a concentration series starting at higher concentrations (~10mg/ml) and going down to smaller concentrations (~1mg/ml). In the end, the highest concentrated sample that shows no aggregation is evaluated.

- Concentration of the samples: concentrations for diluted samples measured by SAXS/SANS are typically situated in the range from 1-10 mg/ml (this corresponds to molarities of 0.05-0.5 mM for a 20 kDa system and 0.01-0.1 mM for a 100 kDa system). An accurate determination of the concentration prior to the experiments is also important for an absolute measurement of the molecular masses in solution by SANS (Zaccai, Jacrot 1981 link to paper if possible). Concentrations should be known at an accuracy of about +/- 10%. In general, optical density measurements (e.g. at 280 nm for proteins) are more accurate than other assays (Bradford etc…).

3) During the experiment: Depending on the size of the macromolecules studied, the sample volume available, the temperature and pressure required, the instrument needs to be set up specifically.

- Angular range: Collimator-sample and detector-sample distances need to be optimized against the size (i.e. presumed radius of gyration) of the macromolecules under investigation. If large objects are studied, these distances will generally be chosen large, if smaller objects and higher angular ranges are required, these distances will be chosen shorter.