Dear Huihun Jung :

DLS can measure the hydrodynamic diameter of your protein or protein aggregate. They should be in Brownian motion in some depresants (buffers). The software you are entering the refractive index and the viscosity depresants. If your protein is not dissolved, then you will not be able to use DLS. Or you need to disperse their proteins to the size of 100-1000 nm.

Best regards.

You can probably learn something about the aggregation of your modified protein, but it may be quite tricky. If the protein is entirely insoluble then it would not be possible to do DLS unless it is dispersed somehow (for example with the help of surfactants). In reality, you may find that depending on your modification you observe different degrees of solubility, and DLS may help discern the level of solubility (average size and average polydispersity), for example versus temperature or when looking at different buffers/additives.

http://www.materials-talks.com/blog/2014/11/17/tips-tricks-for-nanoparticle-characterization/#aggregatingparticles

http://www.materials-talks.com/blog/2016/04/21/aggregation-kinetics-with-dls/

DLS is a technique used for determining the hydrodynamic radius of a particle in solution. In your case the particles are protein molecules. When they are soluble in a buffer we "do not observe scattering" rather they show absorption in the UV region light. However, when protein molecules start to unfold which can be due to different physical (heat, mechanical energy ) or chemical (acid/alkali/chaotropic agents like urea, GdnHCl etc) agents. During unfolding process the inter molecular interaction between protein molecules and solvent is disfavored. As a result the protein-protein interaction is highly favored which results in aggregation of the proteins in the solution. Now we observe them as precipitate or as cloudy suspension. These aggregates have the tendency to scatter light and thus we use DLS for monitoring their aggregation behavior. This is represented in terms of hydrodynamic diameter of the aggregates/microggregates. One more parameter that we get from DLS studies is the population of a particular type aggregate.

Hope it will be helpful...

DLS is not appropriate for particles that are so large they settle out of suspension. Larger particles, up to a few microns, may be investigated using laser diffraction.

http://www.malvern.com/en/products/technology/laser-diffraction/

My protein spectrum is always in aggregates how can i explain tis?

Dynamic light scattering is extremely sensitive to aggregates. If even a small percentage of your protein is in large aggregates, the DLS intensity distribution will appear to be essentially all aggregates. It would be advisable to check by another method, such as gel filtration chromatography, to see if the DLS is exaggerating the problem.

Some DLS software will convert from intensity distribution to mass distribution, using a calculation based on the relationship between particle size and scattering intensity. If the proportion of aggregate is small, this procedure will yield a more accurate result for the protein size. If the protein is mostly aggregated, the mass distribution will also reflect that.

If your protein really is all or mostly aggregated, you will have to revise the method of preparation and/or storage to identify better conditions for obtaining an unaggregated protein.

Hi Bayane Sabbagh , as Adam indicated, this is possibly related to sample preparation, combined with the fact that light scattering is very sensitive to even very few larger components in the sample. You could try filtration (for example with 20nm poresize) or centrifugation, or a different buffer, pH, condition. Lastly, it may help to take lots of very short subruns for the correlation function.