IMPROVED DIRECT PIEZOELECTRIC BIOSENSORS OPERATING IN LIQUID SOLUTIONFOR THE COMPETITIVE LABEL-FREE IMMUNOASSAY OF 2,4-DICHLOROPHENOXYACETIC ACID

The development of a label-free direct piezoelectric immunosensor is r eported. The piezoelectric crystals were modified with 2,4-dichlorophe noxyacetic acid (2,4-D) using coupling procedures based on self-assemb led monolayers of suitable thiocompounds (aminothiophenol, cysteamine, cystine and dithiobis(succinimidyl)propionate) on the surface of gold electrodes of the crystals. The piezoelectric biosensor has been plac ed in a flow-through cell and the affinity binding of two monoclonal a ntibodies against 2,4-D on the modified piezoelectric crystals was stu died in real time without any additional labels. For the interactions, the kinetic equilibrium association constants K-a were 12.6 and 25.0 mu M-1 for the antibodies E2/G2 and E4/C2, respectively. The system wa s used for competitive determination of free 2,4-D in water. For the a ntibodies E2/G2 and E4/C2 (both in concentration of 15 mu g/ml), the l imits of detection (10% decrease of relative binding of the antibody) for free 2,4-D were 0.27 and 0.24 mu g/l, respectively. The total time for one measurement was below 25 min (10 min preincubation of antibod y with sample, 10 min binding reaction, 5 min regeneration).