Photoacoustic imaging offers optical contrast images of human tissue at acoustic resolution, making it valuable for diverse clinical applications. However, quantifying tissue composition via optical contrast remains challenging due to the unknown light fluence within the tissue. Here, we propose a method that leverages known chromophores (e.g., arterial blood) to improve the accuracy of quantitative photoacoustic imaging. By using the optical properties of a known chromophore as a fluence marker and integrating it into the optical inversion process, we can estimate the unknown fluence within the tissue. Experimentally, we demonstrate that this approach successfully recovers both the spectral shape and magnitude of the optical absorption coefficient of an unknown chromophore. Additionally, we show that the fluence marker method enhances conventional optical inversion techniques, specifically (i) a straightforward iterative approach and (ii) a gradient-based method. Our results indicate an improvement in accuracy of up to 24.4% when comparing optical absorption recovery with and without the fluence marker. Finally, we present the method's performance and illustrate its applications in carotid plaque quantification.

Sentinel lymph node (SLN) biopsy is an essential procedure for accurate disease staging and treatment planning in patients with melanoma and breast cancer. Conventional preoperative imaging primarily utilizes lymphoscintigraphy with technetium-99m (Tc-99m), which presents several limitations, including radiation exposure, logistical challenges, and potential delays in surgical workflow. Photoacoustic imaging (PAI) has emerged as a promising alternative, leveraging optical contrast provided by indocyanine green (ICG). A feasibility study was conducted at Erasmus MC, University Medical Center Rotterdam, to assess the potential of dual-wavelength PAI for SLN mapping. PAI was employed to perform spectroscopic measurements in healthy volunteers, supporting the development of an optimal excitation protocol. Subsequently, in the patient phase, SLN mapping was performed using PAI with ICG, and the results were compared to the standard-of-care method utilizing Tc-99m. The excitation wavelengths of 800 nm and 860 nm were selected for ratiometric imaging to effectively visualize ICG while suppressing clutter from hemoglobin and melanin. Among the eleven evaluated sentinel nodes, seven were successfully identified using PAI. The maximum SLN detection depth achieved with PAI was 22 mm. This study illustrates the feasibility of ICG-enhanced dual-wavelength PAI for preoperative SLN mapping in patients with melanoma and breast cancer, as an alternative to lymphoscintigraphy. Analysis of false-negative detections suggests improvements to PAI and optimal patient selection. The proposed ratiometric PAI methodology, compared to multiwavelength spectroscopic imaging, enables faster imaging speeds and facilitates the transition to cheaper light sources.