Screening of small molecule drugs is relevant for toxicological and forensic analysis and immunoassays are predominantly used for this purpose. Enzyme linked immunosorbent assays (ELISA) are the most popular immunoassay method due to their simplicity and low cost. However, reproducibility is problematic, even with automation. In addition, competitive ELISA usually requires the attachment of a larger enzyme to small molecules for detection, which can alter the kinetics of binding. An alternative method for label-free detection of small molecules is second harmonic generation (SHG). SHG is a coherent, second-order nonlinear optical process which has previously been demonstrated for use in immunoassays. Cocaine, methadone, and morphine were used as test cases in a SHG immunoassay. Using detection via SHG in a primary antibody assay, the designed antibodies for these drugs show a high degree of cross-reactivity. Interestingly, the competitive SHG immunoassay showed that specificity of anti-morphine antibodies for morphine could be achieved when a competition assay utilizing the drugs and a labeled drug conjugate were employed, illustrating the kinetic advantage competitive assays provide.

Kinetic advantage of a competitive assay for label-free detection in small-molecule immunoassays

256th National Meeting (2018)

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