Abstract: The present invention includes novel isoelectric focusing cells and methods for isoelectric focusing of amphoteric substances within fluids containing carrier ampholytes which inhibit the phenomena known as spiking or acid notching. The apparatus includes a barrier separating the electrode compartments from the focusing cell compartments which minimizes hydrogen and hydroxyl ions contained in electrolyte solution within the electrode chambers from passing into the focusing cell passageways. The currently preferred barrier is a bipolar membrane; however, the combination of anion and cation selective membranes is also effective. Such a barrier enables very narrow pH gradients to be established and maintained, thereby permitting high resolution separations of amphoteric biological substances.

Abstract: The present invention includes novel apparatus and methods for isoelectric focusing of amphoteric substances within fluids containing carrier ampholytes. The present invention utilizes multichannel recycling isoelectric focusing techniques which eliminate mixing of fluid which passes through the isoelectric focusing cell with the original crude fluid sample. Mixing is avoided by preferably using a dual reservoir system and alternating the fluid flow into and out of each reservoir after each pass through the isoelectric focusing cell. Fluid flow is alternated into and out of each reservoir of each reservoir pair through use of reservoir inlet and outlet valves. The reservoir inlet and outlet valves preferably operate in response to the fluid level within the reservoir supplying fluid to the isoelectric focusing cell. When the reservoir empties the corresponding reservoir outlet valve automatically engages to stop fluid flow from the empty reservoir and to start fluid flow from the full reservoir.