FIGURE 6. Quantification of a competitive immunoassay for phenytoin using surface plasmon resonance (SPR).

a, SPR difference image used to quantify a competitive immunoassay for phenytoin (manuscript in preparation, K.N. et al.). The detection zone is a gold surface precoated with bovine serum albumin (BSA) covalently modified by phenytoin. The three streams (750 nl s^-1) contained 150 nM anti-phenytoin antibody mixed with 0 nM, 50 nM or 100 nM soluble phenytoin (bottom to top, respectively). The contrast in the image has been adjusted to highlight the differences between the nonfouling polyethylene glycol (PEG) upstream and the three sample regions. b, c, Competition assay detection of low-end therapeutic levels of phenytoin in a model system (b) and in preconditioned saliva (c). b, SPR reflectivity over time of anti-phenytion antibody in phosphate buffer binding to a BSA–phenytoin-treated surface. The plot shows that the rate of antibody binding negatively correlates with the amount of competitor (phenytoin) added to the solution. c, SPR reflectivity of variously treated preconditioned saliva samples over time. In this case, whole human saliva was preconditioned using a mechanical filter and the H-filter off-card. Plots b and c show that the PEG region effectively resisted fouling to either antibody or components present in preconditioned saliva.