Multi-Angle Light Scattering (MALS) measures the amount of light scattered by particles in solution relative to the angle of the incident light. For larger macromolecules and complexes (above 10 nm for 690 nm light) or for unfolded or strongly elongated proteins, the angular dependence in a multi-angle light scattering experiment can be used to calculate Rg. Rg is the “root means square radius” or “radius of gyration” and reflects the mass distribution of a macromolecule around its center of mass.

MALS in conjunction with a concertation concentration detector can directly measure the absolute molar masses of macromolecules and complexes, independent of structure and shape from 200 daltons to hundreds of millions of daltons. SEC-MALS uses size exclusion chromatography (SEC) coupled to an in-line multi- angle light scattering detector and an in-line refractive index or UV/Vis detector. SEC-MALS can be used to determine the absolute molecular mass of proteins and complexes in complex samples, independent of their elution volume from the SEC column. In contrast to conventional SEC chromatography this method can accurately measure MW, even for sticky or elongated proteins that elute to late or too early for their actual mass to be correctly determined by size exclusion chromatographySEC alone (due to their unusual interaction with the columns).

An integrated dynamic light scattering detector allows measurement of the hydrodynamic radius (Rh)  of particles, with a lower limit of 0.5 nm.

In CG-MALS a composition-gradient preparative system is combined with MALS/RI/UV detectors to measure interactions between biological macromolecules, by directly observing changes in molar mass due to complex formation or dissociation in solution and without the need for labeling or immobilization.

 

MALS Instruments

We have two SEC-MALS Systems; both MALS/RI detectors can also be used for batch measurements.
  • Wyatt Dawn EOS 18-angle light scattering detector combined in line with Wyatt Optilab DSP refractive index monitor linked to a Shimadzu HPLC LC-20AD pump. An additional UV detector can be added to the system.
  • Wyatte Dawn Helleos II 18-angle light scattering detector in-line with a Wyatt Optilab rEX Refective index detecor linked to an AKTA purifyer system for SEC and UV monitor.

Accessories:
Calypso, a composite gradient preparation system that can be combined with the  Dawn Helleos II 18-angle light scattering detector, the Optilab rEX Refractive Index detector, and the AKTA purifier UV monitor for CG-MALS experiments.

QELS detector integrated in the Dawn Helleos II for simultaneous DLS measurements

Comet cell cleaner for the Dawn Helleos II

Orbit Solvent Recycler integrated with the Dawn Helleos II set up

Sec-MALSGC-MALS

Exciting Possibilities in Biology, Biochemistry and Biophysics:
  • Absolute molar mass of macromolecules and complexes  
  • Absolute molecular stoichiometry
  • Binding affinity from pM to mM
  • Self- and hetero-association and simultaneous self- and hetero-association
  • Second virial coefficient A2
  • Molar Mass and  stoichiometry of membrane protein-detergent complexes
  • Protein-conjugate analyses ( e.g. glycosylated or pegylated proteins)
  • Non-specific interactions between proteins at high concentrations
  • And a lot more ...

SEC-MALS and CG-MALS example data

Users have to receive hands-on training, from Dr.Erbse, before they are allowed to use the instrument independently. Interested users should contact Dr. Erbse (details below) to schedule training. The training will cover sample preparation, performing measurements, cleaning the SEC-MALS/CG-MALS set-up and data evaluation. Training will be done employing a standard sample in addition to actual samples provided by the user.

The protocols give step-by-step instructions for a basic SEC-MALS or CG-MALS experiments, assuming samples have been prepared correctly and the user has been trained on the instrument. Reading the protocols DOES NOT replace the TRAINING provided by Dr.Erbse.
 

After your training is completed, you will be invited to join the MALS Google Calendar. To reserve MALS time, sign up with your name and lab. Please be sure to avoid booking time that overlaps with other users.

One SEC-MALS is located on the third floor of JSCBB in the D-Wing, room D381, on East Campus. Proxcard access is required after normal hours and on weekends. The second SEC-MALS and the CG-MALS are located in the Luger groups laboratory (second floor A wing of the JSCBB,  A wing room A255). After hour and weekend access MUST be coordinated with the Luger lab.

  • Users are expected to provide all consumables specific to their experiment
  • Users are expected to provide their own SEC columns.
  • Upon request it might be possible to borrow columns from other user groups. Users are expected to replace any borrowed columns that were damaged by them
     
    • SEC columns for Light Scattering MUST be in top condition and should not "bleed" any column material.
    • Most columns used frequently for "normal" gel filtration application will be unsuitable for SEC-MALS
    • Users are expected to pay for repairs or parts that are necessary due to damage caused by their carelessness or neglect.
       
  • Cost for repairs/parts/services resulting from normal wear will be split based on the time used between all user groups.
  • Please Note -
    Since repairs/service from normal wear might not be necessary every year, depending on use, these costs might come quite some time after use.