LOW-COST FLOW-THROUGH ISOELECTRIC FOCUSING

THIS PROPOSAL WILL EVALUATE AND DEFINE THE USEFULNESS OF A NOVEL APPROACH FOR THE SEPARATION AND PURIFICATION OF BIOLOGICAL COMPONENTS. THIS APPROACH WILL BE A MODIFICATIONOF THE ISOELECTRIC FOCUSING (IEF) PROCEDURE. THE MODIFICATION UTILIZES PH GRADIENTS ESTABLISHED BY DIFFUSION ACROSS DIALYSIS MEMBRANES WHICH SEPARATE THE AMPHOLYTES FROMTHE SPECIES BEING FOCUSED. THE PROCEDURE HAS SEVERAL ADVANTAGES OVER EXISTING TECHNIQUES: IT HAS HIGH CAPACITY AND CAN BE USED IN A CONTINUOUS FLOW-THROUGH FASHION; IT IS LESS EXPENSIVE THAN OTHER IEF TECHNIQUES; AND THE PH GRADIENT MAY BE ALTERED DURING THE FOCUSING RUN TO ACHIEVE MAXIMAL SEPARATION OF ONE OR MORE COMPONENTS FROM A GIVEN STARTING MATERIAL. THE SPECIFIC AIMS OF THE PROPOSED RESEARCH ARE: (1) TO INVESTIGATE WAYS TO DECREASE THE RESOLUTION TIME OF THE DEVICE, (2) TO STUDY THE DEGREE TO WHICH THE PH GRADIENTS MAY BE ALTERED DURING THE RUN BY THE ADDITION OF AMPHOLYTES OF SELECTED RANGES, AND (3) TO INVESTIGATE THE LOADING CAPACITY OF THE SYSTEM.