Practical Aspects of Multiwavelength Analytical Ultracentrifugation

Abstract

Open-source Multiwavelength Analytical Ultracentrifugation (MWL-AUC) detection systems have been evolving for over a decade. Continual advances emerging out of several research groups have brought the instrumentation technology to increasingly higher levels of performance. The capabilities of MWL-AUC have been documented in many publications, demonstrating the applicability of broad spectrum absorbance acquisitions in analytical ultracentrifugation to a wide array of scientific fields. Despite numerous examples of the usefulness and unique advantages of MWL-AUC, the adoption of the technology by more research groups has been slow. The complexity of the hardware, integration within an ultracentrifuge platform and lack of practical construction and operational information is the likely source of reluctance. Here, we clearly describe the challenges facing a researcher considering adopting MWL-AUC technology in their own laboratories, and provide the information necessary to implement and operate a MWL-AUC system. The discussion includes details of detector assembly, optical alignment, and acquisition parameter settings necessary to achieve high quality experimental results.

A multiwavelength emission detector for analytical ultracentrifugation

“…The MWE-AUC design, as shown in Fig. 2, is based partly on concepts used for MWL-AUC 31 and the established confocal fluorescence detector. 12 There are two main setup parts: the scanning unit and the signal-coupling unit.…”

“… 22 Details on the AUC triggering have been recently published elsewhere. 31 In short, the temporal delay between the detection of a polished area on the rotor bottom via a light barrier and the sample cell arriving at the measurement position is a function of rotor speed and has to be calibrated using a fluorescent dye solution as a test sample.…”