Visualization of BChE bound to immobilized monoclonal antibodies

To confirm that the BChE in plasma had been captured by immobilized antibodies rather than simply inactivated, we extracted bound protein from the beads and visualized the extracted proteins on an SDS slab gel stained with Coomassie blue. In brief, beads were transferred to 0.45 micron spin filters and washed with 0.5 mL aliquots of phosphate buffered saline until the absorbance at 280 nm of the flow through was <0.04. The washed beads were desalted by two washes with 0.5 mL water. Bound proteins were released from the antibody by two treatments with 50 μL of 1% trifluoroacetic acid. The extracts were dried in a vacuum centrifuge, dissolved in 20 μL of SDS/dithiothreitol, glycerol, bromphenol blue, denatured in a boiling water bath for 3 min and loaded on an SDS gel.