Multi-wavelength absorbance optics The multi-wavelength system provides an absorbance spectrum at each radial position (Strauss et al. 2008; Bhattacharyya et al. 2006). Deconvolution of these spectra allows the discrimination of individual species sedimenting in complex mixtures of molecules. Cellular and molecular biology will benefit from this optical system since it will allow the size distributions of labeled molecules to be determined in a complex milieu (e.g. cell lysates). Molecules with different chromophores, such as DNA binding proteins, heme proteins or ligands and tagged molecules can be analyzed to study assemblies and complexes. For example, it will be possible to identify the components and characterize the cellular conditions under which molecules sediment as part of a macromolecular complex. Figure 3 shows the current multiwavelength detector and also the sedimentation of bovine serum albumin detected by two different operation modes: the time mode which detects the time dependent sedimentation of the sample with the detector set at a fixed radius and the radial mode which scans the cell radius at a given time. The optics will use a constant light source, a rapid scanning stepping motor, and a fast Andor ICCD camera capable of nanosecond integration time, allowing signal collection of all cell channels in each rotor revolution even at speeds as high as 60,000 rpm.
Fig. 3 The multiwavelength detector arm mounted in an XL-A AUC (left) as well as the typical experimental traces in time mode with radially fixed detector and radial mode scanning the AUC cell radially at a fixed time. The x-axis in these plots is the wavelength and the z-axis absorption. The data shown are for sedimenting bovine serum albumin (BSA)