Concentration and fractionation by isoelectric trapping in a micropreparative‐scale multicompartmental electrolyzer having orthogonal pH gradients. Part 2

Abstract

A micropreparative-scale multicompartmental electrolyzer called ConFrac has been developed and tested for isoelectric trapping separations. ConFrac can be operated in pass-by-pass mode or recirculating mode, using either asymmetrical feeding (feed enters only the anodic or the cathodic flow-through compartment) or symmetrical feeding (feed enters both the anodic and the cathodic flow-through compartment). Symmetrical feeding results in higher processing rates and is the preferred operation mode. Residence time in the flow-through compartments is set as a compromise between processing rate and temperature rise in the effluent. Ampholytic components have been isolated from 5 to 50 mL volumes of micromolar feed solutions and hundredfold concentrated into 100-μL collection compartments. Samples containing ampholytic analytes in highly conducting salt solutions were readily desalted and fractionated in ConFrac in one operation. pH transients formerly observed in other isoelectric trapping devices were observed in ConFrac as well. The pH transients were caused by the unequal ion transmission rates of the anodic- and cathodic-side buffering membranes.

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.…”

A novel microscale preparative gel electrophoresis system

“…It was a novel idea but not suitable for practical samples. Lim et al 19 designed an isoelectric trapping apparatus in the design called concentrator-fractionator. The apparatus was able to isolate ampholytic components from recirculated feed solutions.…”

Preparative‐scale isoelectric trapping by recursive electrophoresis in a compartmentalized system having orthogonal primary and secondary pH gradients. Part 1 – Construction and standard operation

“…Though this system has been applied to a number of protein samples, it can only process microgram quantities of proteins per gel plug [24]. Another parallel-arranged system, the ConFrac [25][26][27], has been developed to process milligram quantities of ampholytes present in o10 mL of solution. The ConFrac contains several minimalistic IET heads that are placed side by side.…”