Down - Light scattering

Question 1

[Light scattering] Why do we use light scattering? (6)

Answer 1

Everything scatters light – no need for absorbance, fluorescence or chromophore

Get functional mass and complex formation under native conditions (solution)

No upper limit (as MS) (the larger the molecule, the lower the detection limit)

Large enough molecules (approach wavelength) > size (radius of gyration) of the molecules

Real time method, follow kinetics of degradation/synthesis or disassociation/association

Get diffusion coefficient

Question 2

[Light scattering] How do we get molecular mass from LS?

Answer 2

Relation through relative intensity of scattering (with certain angle between insicdent and observed light).

“By means of intensity between scattered light and incident light we can measure the molar mass”

Question 3

[Light scattering] How does Dynamic Light Scattering work?

Answer 3

Fluctuations in scattering intensity can be translated to a diffusion constant, into radius of gyration. Smaller particles move around/diffuse more = more fluctuations.

Question 4

[Light scattering] What can a change in radius of gyration indicate?

Answer 4

Denaturation of proteins, or other forms of DNA (linear rod or circular plasmid).

Use large angles to differentiate between forms with different radius of gyration.