SELECTIVE PRECONCENTRATION FOR CAPILLARY ZONE ELECTROPHORESIS USING PROTEIN-G IMMUNOAFFINITY CAPILLARY CHROMATOGRAPHY

Capillaries with 150 mu m inner diameter were packed with a perfused p rotein G chromatographic support and used as immunoaffinity preconcent rators for capillary zone electrophoresis. Antibody was loaded onto th e protein G support to form an immunoaffinity stationary phase. Inject ion of samples onto the column caused selective retention and preconce ntration of antigen. Injection of appropriate buffers onto the column caused desorption of the antibody and antigen which were then separate d by capillary zone electrophoresis. The combination was used on-line and off-line. For on-line combination, a flow-gated interface coupled the two columns and allowed injection of desorbed zones onto the elect rophoresis system. Off-line coupling required collection of desorbed f ractions and then injection onto the electrophoresis system. Flow rate s as high as 100 mu L/min were used to load sample onto the affinity c olumn. Desorbing flow rates had to be 1 mu L/min or less to prevent ex cessive dilution during desorption. Using the system, 1 mt insulin sam ples could be loaded onto the affinity column and desorbed in volumes as small as 1 mu L for 1000-fold preconcentration. The use of the prec oncentrator with serum samples spiked with insulin was demonstrated.