Measurement of fluorophore lifetimes provides a sensitive probe of the microenvironment of the fluorophore, with various applications in biology and medicine. Its sensitivity is not affected by the concentration of the fluorophore or by the excitation light intensity. This is particularly significant in biological and clinical applications where dye concentration cannot be precisely controlled and light fluence is highly non-uniform. Optical imaging techniques have been developed to map the distribution of fluorescence lifetime in a wide field of view, and have been applied for functional imaging of cellular metabolism, oxygen sensing, mapping the concentration and dynamics of ions, and for probing molecular associations by sensing intra-molecular distances. In all these applications the lifetime is evaluated by measuring the decay of the fluorescent signal, or by transient absorption. One of the major difficulties in applying these methods for clinical imaging is the loss of spatial information due to strong light scattering.
Researchers at the University of Michigan have developed a method to locally measure a dye’s lifetime in the excited state. This is accomplished via photoacoustic probing, whereby the dye an is excited by an excitation laser pulse and probing pulse is used to generate photoacoustic waves, which decay, resulting in an intensity point. The process can be repeated, to determine the slope between the first and second intensity points. This method has been demonstrated for oxygen level measurement using oxygen sensitive dye and also for pH imaging using ratiometric photoacoustic chemical sensing, with further application in in vivo imaging to provide image contrast. The agents act as photoacoustic sources, changing their signal generation efficiency as a function of the specific external parameter, thus providing an image of the distribution of biologically relevant parameters in living tissue.
Applications and Advantages
- Medical imaging diagnostics and therapy, including non-invasive, clinical functional imaging.
- In vivo mapping molecular events such as resonant energy transfer and ion collisions
- Does not require collection of emitted light
- Provides deep tissue imaging and optical contrast