AUC 2022

Workshop on High Concentration FDS-AUC 1)


Presenters: Jack Correia, University of Mississippi Medical Center, Jackson, MS, USA, and Walter Stafford, Harvard Medical School, Cambridge, MA, USA.


Since the early days of protein characterization, the development of AUC analysis for high concentration SV data has been a critical focus. The early Model E Schlieren optical system worked best from 5-20 mg/ml. With the advent of therapeutic antibodies delivered at up to 150 mg/ml, high concentration AUC techniques continue to be proven irreplaceable. This workshop will discuss two essential steps in FDS AUC data collection and analysis:

  1. Experimental setup of FDS SV runs at high concentrations.
  2. Analysis of FDS SV data with nonideal, associating models in SEDANAL.

We typically perform experiments in tracer mode with 100 nM Alexa-labeled mAbs in unlabeled mAb concentrations from 1 to 150 mg/ml. FDS optics do not need reference channels, so we run up to six samples in a four hole rotor with 3 mm centerpieces. For this reason, sample labeling and equilibration will be discussed. We process FDS data for meniscus and base regions, and globally fit with models that account for hydrodynamic k s and thermodynamic nonideality BM 1 , plus weak association. Therefore, basic SEDANAL procedures (data preprocessor and ModelEditor) will be presented. Lastly, we perform error analysis by bootstrap with replacement methods. Experiments can also be performed in human serum, which necessitates knowing serum protein concentrations and the use of matrix methods for k s and BM 1 cross term nonideality parameters. Data sets will be provided for student testing. SEDANAL runs in Windows 10 or 11, and on a Mac, requires Parallels, Boot Camp or VWware. Students should prepare for this workshop by reading Correia, et al. Eur. Biophys. J 49, 687, 2020.

Please note:
The current version of SEDANAL along with the User Manual can be downloaded from

This workshop will be presented on Sunday from 14:30-16:30 (session 3).