Non-invasive quantitative acoustic imaging of intact brain organoids for structural assessment in drug discovery

Quantitative characterization of microstructural variations in human brain organoids is essential for advancing both fundamental neuroscience and translational applications, such as drug discovery and understanding tissue development and pathology. In this study, intact human brain organoids were developed and imaged using high-frequency Scanning Acoustic Microscopy (SAM) with 40 MHz and 60 MHz transducers. Both transducers were experimentally characterized and numerically simulated to assess their performance and confirm their suitability for high-resolution acoustic imaging. The statistical distribution of the scattering amplitude was analyzed using the Rayleigh and Nakagami models to infer tissue heterogeneity. Multiple regions of interest (ROIs) within the organoid structures were examined, revealing that the Nakagami distribution–with its additional shape parameter–provided a better fit to experimental data, capturing subtle microstructural differences not evident in conventional B-mode images. Across all ROIs, the Nakagami shape parameter m ranged from 0.62 to 0.99, corresponding to uniform neuroproliferative zones and heterogeneous differentiated regions, respectively, demonstrating the method’s ability to distinguish developmental compartments. These findings demonstrate that statistical acoustic imaging, supported by transducer performance validation, provides a non-invasive and quantitative method for examining brain organoid architecture, facilitating enhanced evaluation of tissue response in preclinical drug development and studies of tissue growth and disease progression.