Non-resonant multiphoton photoacoustic spectroscopy for noninvasive subsurface chemical diagnostics

This paper describes the development and validation of a novel noninvasive spectroscopic subsurface chemical detection technique, non-resonant multiphoton photoacoustic spectroscopy (NMPPAS). In this technique, non-resonant multiphoton excitation is used to provide subsurface excitation of chemical constituents in a sample followed by the subsequent detection of an acoustic signal using a piezoelectric transducer. Because NMPPAS relies on non-radiative relaxation of the absorbing species, it is capable of monitoring both fluorescent and non-fluorescent species. Moreover, since the majority of the energy imparted to most molecules upon the absorption of light is released through non-radiative pathways, sensitive measurements of even fluorescent molecules can be performed. In this paper, demonstration of proof-of-principle of this novel technique has been shown using test samples of common fluorescent dyes and biomarkers including rhodamine 6G, tryptophan, and NADH in solution and gelatin tissue phantoms. From these studies, it was found that detection limits of these chromophores are in the subnanomolar concentration regime. In addition, preliminary results on excised tumor and healthy tissue samples have demonstrated significant differences between the tumorous and non-tumorous tissues at 740 nm and 950 nm wavelengths. From this work, it was found that NMPPAS has a great deal of potential for subsurface chemical diagnostics in the field of biomedical research.