Surface-enhanced Raman spectroscopy combined with atomic force microscopy for ultrasensitive detection of thrombin

We have developed an ultrasensitive analytical method based on surface-enhanced Raman spectroscopy (SERS) exploiting a Raman probe covalently bound to gold nanoparticles. The biological marker to be detected was adsorbed on functionalized gold nanoparticles. The capture of these nanoparticles via a biorecognition process between the marker and the immobilized receptor was demonstrated by atomic force microscopy (AFM) imaging. The vibrational fingerprints of the Raman probe on the capture substrate were followed to reveal the presence of the biological marker. The method, which was applied to reveal thrombin captured on a substrate containing antithrombin and heparin, resulted in the ability to detect marker concentrations down to the picomolar (pM) level.