Preparative‐scale isoelectric trapping by recursive electrophoresis in a compartmentalized system having orthogonal primary and secondary pH gradients. Part 2 – Operation in cascade mode

Abstract

A parallel multicompartmental electrolyzer recently developed for preparative-scale isoelectric trapping separations, trapping by recursive electrophoresis in a compartmentalized system, was set up to operate as a cascade of binary separations to produce at least one pure target ampholyte (or more, with additional separation heads) without other ampholytes ever entering (even transiently) the harvest stream. This mode of operation avoids the need for exhaustive electrophoresis and the accompanying long separation times brought about by the exponentially decreasing concentrations over the course of batch separations. Continuous operation can be achieved in the cascade mode by continuously feeding the sample into the first separation head configured with three flow-through compartments and continuously harvesting one (or more) target components in additional separation heads configured with two flow-through compartments, attached to the respective branching points.

Isoelectric Point Separations of Peptides and Proteins

“…Various reports have highlighted the advantages of the MSWIFT device including desalting procedures, separation of small and large ampholytic components [54], bottom-up proteomic studies [55,56], and unique applications for purification [57]. Other IET devices have been constructed including the ConFrac [58,59] and TRECS [60,61] systems. It should be noted that any MCE that utilizes buffering membranes can be operated in IET mode.…”

IEF‐based multidimensional applications in proteomics: Toward higher resolution

“…Parallel IEF reported recently, in reality, does not represent a real IEF, but rather can be classified as a dynamic trap . Another interesting idea is a preparative‐scale isoelectric trapping by recursive electrophoresis in a compartmentalized system .…”

Preparative isoelectric focusing in a cellulose‐based separation medium

“…IEF is a particularly effective way of the separation of ampholytic compounds [2,3]. Although IEF is usually performed in a gel, several other IEF systems are available for separation of ampholytic compounds [2][3][4][5][6][7][8][9][10]. Preparative IEF using a Sephadex gel was introduced by Radola [11] and more recent use of this method was reported by Görg et al [12].…”