SLS - Static Light Scattering.
SLS measures the time-averaged intensity of light scattered by particles in solution.
When to use SLS.
Unlike DLS, which analyzes fluctuations in scattered light over time, SLS measures the total amount of light scattered. This provides information about molecular weight, protein-protein interactions, and overall sample quality.
SLS is particularly valuable for therapeutic protein development, where high-concentration formulations are often required. B22 measurements can predict whether a candidate will remain stable and low-viscosity at clinically relevant concentrations.
SLS is ideal when you need to:
The science behind SLS.
Scattering intensity depends on molecular weight
When light passes through a solution containing macromolecules, some light is scattered at angles away from the incident beam. The intensity of scattered light depends on the size and concentration of the scattering particles.
Larger molecules scatter more light than smaller molecules. By measuring the scattered light intensity at a fixed angle and knowing the sample concentration, you can calculate the weight-average molecular weight (MW) of the particles in solution.
The Debye equation relates scattering to molecular properties
SLS measurements are typically performed at multiple concentrations. The relationship between scattering and concentration is described by the Debye equation:
Where:
– K is an optical constant (depends on wavelength and refractive index)
– c is the concentration
– Rθ is the Rayleigh ratio (proportional to scattered intensity)
– Â MW is the weight-average molecular weight
– A2 (or B22) is the second virial coefficient
Plotting Kc/Rθ versus concentration produces a linear relationship. The y-intercept gives 1/ MW, and the slope gives 2*B22.
The second virial coefficient reveals protein-protein interactions
- Positive B22: Proteins repel each other (favorable for high-concentration formulations)
- Negative B22: Proteins attract each other (risk of aggregation or high viscosity)
- B22 near zero: Proteins behave nearly ideally (minimal interactions)
Negative B22 values are a red flag for therapeutic development because they predict problems at high concentrations: aggregation, precipitation, or unacceptably high viscosity.
Parameters you measure with SLS.
Molecular weight (MW)
The weight-average molecular weight of particles in solution. This confirms the oligomeric state (monomer, dimer, tetramer, etc.) or detects aggregation. For example, if you expect a 150 kDa monomer but measure 300 kDa, your protein is forming dimers.
Second virial coefficient (B22)
A measure of protein-protein interactions. Positive values indicate repulsive interactions (good for formulation). Negative values indicate attractive interactions (risk of aggregation).
Initial scattering intensity
A quick quality check. Very high scattering suggests large aggregates or particulates. Very low scattering may indicate the sample is too dilute for reliable DLS measurements.
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