Characteristics of a label-free piezoelectric immunosensor detecting Pseudomonas aeruginosa

A label-free immunosensor system detecting a psychrophylic bacterium, Pseudomonas aeruginosa was developed as follows. Four types of anti-P. aeruginosa antibody were individually chemisorbed onto one-side gold electrodes of piezoelectric quartz crystals according to a thiolated antibody coupling procedure initiated with a thiol-cleavable heterobifunctional cross-linker, sulfosuccinimidyl-6-3-(2-pyridyldithio)propionamido]hexanoate. A flow-type biosensor system was operated optimally at 0.2 M sodium potassium phosphate, pH 7.2 with a minimal matrix effect and the selected flow rate for it was 0.155 ml/min. A biosensor response was detected by measuring a steady-state resonant frequency shift after the response time around 8 min. The frequency shifts obtained were quite specific according to the antibody types and P. aeruginosa strains. The biosensor responses to varying concentrations of the P. aeruginosa cells ranging from 1.3×10⁷ to 1.3×10⁸ CFU/ml were determined as 17–176 Hz and a linear calibration curve (r=0.942) was obtained by plotting the responses in a double-logarithmic scale. The selectivity of the biosensor between P. aeruginosa and Xanthomonas spp. which both belong to the aerobic pseudomonads was, however, not so good owing to the property of the antibody used. The sensor chip could be reused at least seven times without an appreciable decrease in sensitivity.