Molecular specificity in photoacoustic microscopy by time-resolved transient absorption

Area of Science:

• Biomedical Optics
• Molecular Spectroscopy
• Photoacoustic Imaging

Background:

• Traditional photoacoustic microscopy axial resolution is limited by detector frequency bandwidth.
• Transient absorption, a resonant two-photon process, offers a novel approach to enhance resolution.
• Molecular systems with overlapping absorption spectra are challenging to differentiate using conventional methods.

Purpose of the Study:

• To develop and validate a transient absorption ultrasonic microscopy (TAUM) system for ultrahigh resolution imaging.
• To leverage molecular dynamics, specifically ground state recovery times, for differentiating molecular species.
• To demonstrate TAUM's capability in distinguishing closely related molecules, such as oxygenated and deoxygenated hemoglobin.

Main Methods:

• Harnessing transient absorption via a resonant two-photon process for photoacoustic microscopy.
• Reconfiguring a TAUM system with two precisely controlled laser pulse trains.
• Measuring exponential decay time constants related to molecular dynamics, including nonradiative decay.

Main Results:

• Achieved nearly an order of magnitude improvement in axial resolution compared to traditional photoacoustic microscopy.
• Measured ground state recovery time for Rhodamine 6G (3.3±0.7 ns), consistent with its fluorescence lifetime (4.11±0.05 ns).
• Differentiated between oxy-hemoglobin (3.7±0.8 ns) and deoxy-hemoglobin (7.9±1.0 ns) in bovine whole blood.

Conclusions:

• TAUM provides ultrahigh axial resolution, optically constrained by the two-photon process.
• The measured time constant, analogous to fluorescence lifetime, allows differentiation of molecular systems with overlapping absorption spectra.
• TAUM successfully distinguishes between closely related molecular pairs, demonstrating its potential in various scientific applications.