Advances in the measurement of protein mobility using laser Doppler electrophoresis – the <i>diffusion barrier</i> technique

Abstract

A new technique for the measurement of protein mobility using laser Doppler electrophoresis (LDE) is introduced and characterised. The diffusion barrier approach loads a tiny protein sample volume into a much larger volume of dispersant, which contains the electrodes; the LDE measurement is then recorded before the sample can diffuse to the electrodes. We demonstrate that sample volumes are reduced by up to two orders of magnitude to volumes typically associated with separation techniques (∼50 μL), no reduction in measurement sensitivity occurs, samples can be retrieved usefully intact, post-measurement and typical measurement times are of the order of minutes. Measurements of BSA mobility up to 75°C and 1 M buffer concentration and lysozyme at a concentration as low as 0.5 mg/mL are demonstrated using the technique with good agreement with literature values.