Sensitive Analysis of Peptides Using Nonlinear Multi-Photon Laser Wave Mixing and Capillary Electrophoresis

Nonlinear multi-photon laser wave-mixing spectroscopy is presented as a highly sensitive absorption-based detection method for biomedical applications including early diagnosis of neurodegenerative diseases and viral infections.  We are studying wave mixing as a sensitive detector for β-amyloid (Aβ) in order to demonstrate early-stage Alzheimer’s detection.  Wave mixing offers inherent advantages over other methods including parts-per-quadrillion or zeptomole-level detection sensitivity, small sample requirements, compact portable designs, and high spatial resolution that is suitable for protein analysis in a single cell.  The wave-mixing signal is a coherent laser-like beam, and hence, it can be collected with virtually 100% efficiency and minimal background.  The signal has a cubic dependence on laser power and a quadratic dependence on analyte concentration.  In a wave-mixing setup, two laser beams are focused and mixed inside a capillary flow cell.  Capillary electrophoresis is used to separate and identify the peptides.  Wave mixing detection sensitivity levels are comparable or better than those of popular fluorescence detection methods, and yet wave mixing can detect biomolecules in their native form without using labels since it is an absorption-based method.  To natively detect β-amyloid, the amino acids, tyrosine and phenylalanine, are probed with a 266 nm laser.  Wave mixing yields excellent detection sensitivity levels even when using thin (micrometer) analytes.  Hence, it is inherently suitable for interfacing microfluidics, microarrays and capillary electrophoresis separation systems to enhance chemical specificity levels.  Potential applications include early disease diagnosis, sensitive detection of biomarkers and bio agents, cellular protein studies and protein interaction studies.