An apparatus for membrane-confined analytical electrophoresis

A membrane-confined analytical electrophoresis apparatus for measuring the solution charge of macromolecules has been described previously (T. M. Laue et al., Anal. Biochem. 1989, 182, 377-382). Presented here is a design for this apparatus, which permits the on-line acquisition and display of absorbance data from up to 512 positions along an analysis chamber. Concentration distributions of macromolecules in solution can be monitored in the chamber to provide steady-state electrophoresis, electrophoretic mobility and diffusion measurements. Buffer chambers press semipermeable membranes against the open ends of a fused-silica cuvette to form the analysis chamber. This configuration permits both the flow of buffer and the establishment of an electric field across the cuvette, while retaining macromolecules in the field of view. Though a gel may be included in the analysis chamber, none is required for gradient stabilization. The volume of sample required for analysis is 8 microL, most of which is recoverable. Experimental conditions can be varied during study by simply changing the circulating buffer and/or the electric field. The analysis and buffer chambers are held in an aluminum housing that sits in an aluminum water jacket. The water jacket provides temperature control, shielding from external electrical noise and also serves as an optical mask. Plans for the cell assembly, optical system and the computer interface for data acquisition are provided. The assembly and operation of the apparatus and the analysis of data are described.