RESOURCES

Name of Method: MALLS (Static light scattering, SLS)

Result expected: The measured molecular mass of the protein complex to an accuracy of ~2-3%

K_D range: N/A - complex must be stable under purification conditions.

MW range: Current range of measurements performed is 7 kDa - >600 kDa

Purity: High purity required

Labeling-Tagging: No labeling or tagging needed.

Accuracy: 2-3% accuracy of overall molecular weight.

Quantity required: For a complex AB, assuming that A is the 'receptor' and B the 'ligand' how much protein is needed to characterize the complex fully (e.g. obtain K_D or get MW)?
A total of 200 µl of AB at a concentration of 1 mg/ml, total of 0.2 mg of AB.

• N.B. larger particles scatter more, 100µg is sufficient for a 200 kDa protein

HTP: Is the method HTP (can it use 96-well sample plates in an automated fashion)?
No

Speed: Will the user have a quantitative answer immediately after collecting the data?
Processing takes 5 minutes

Cost: (approximate cost of consumables per ONE experiment)

Access: Is an instrument available (name all SPINE2 sites you know)?
Hamburg, Amsterdam.

User expertise: Can the method be provided as a service (measurement and data analysis done by local staff) or only access and assistance on a collaborative basis?
Extremely simple technique, can be performed by inexperienced researcher after 15 minutes tuition; available on a collaborative basis.

Special requirements: Any special sample needs?
Sample must not absorb at 635nm, must be stable on gel filtration for duration of experiment, should be clean (0.2?m filtered).

Method: MALLS

Purpose: MW determination

Range: 3 kDa - 6 MDa, depending on the size exclusion column setup. Three analytical size exclusion columns are available: Superdex 75, Superdex 200 and Superose 6

Accuracy: 2-5%

Sample quantity: 100 ul of 1-10 mg/ml (needed connentration in inverse proportion to sample MW). For example, to analyze a sample of 10 kDa, ~ 8 mg/ml concentration; to analyze as sample of 100 kDa, 1 mg/ml could be enough. Volume of 100 ul is required for one experiment.

Sample purity: should be possible to separate fraction for the analysis from the other fractions of the sample using size exclusion chromatography. If several different types of molecules are present in single chromatography pick the calculated mass will be the average for these molecules.

Speed: 1 hour for the experiment. 4 hours including equilibration of the system before and washing after the experiment.

Special requirements: the system is extremely sensitive to aggregates and small partials. Samples should be filtered through 0.2 um. Samples and buffers should not contain detergents

Access: The NKI system is available to the SPINE-2 collaborators to perform experiments on request.