Ultraviolet photoacoustic microscopy (UV-PAM) provides label-free imaging of cell nuclei with the superior optical absorption of DNAs and RNAs. UV-PAM does not require any additional staining process to visualize the structures of tissues and cells. However, conventional UV-PAM is limited in imaging speed to be used in real pre-clinical and clinical situations. To overcome this limitation, we developed an UV-PAM adopting a high-speed scanning module. We demonstrated the applicability of the system as an intraoperative surgical margin assessment tool by imaging formalinfixed paraffine-embedded (FFPE) sections of human colon cancer tissues and comparing these results to conventional optical microscopic images. Additionally, three more types of cancerous tissue such as liver, uterus, and kidney were photoacoustically examined. The imaging results successfully showed the characteristic features of cancerous and normal tissues, allowing rapid diagnosis of cancer. UV-PAM has achieved sufficient spatial resolution to distinguish the individual nuclei in human tissue, but it is challenging to resolve the closely adjacent nuclei. Therefore, we applied tissue expansion technology, which enables nanoscale imaging of subcellular components beyond the resolution of optical microscopy, to our developed system. A mouse brain section was physically expanded by hydrogel-tissue hybridization, and consequently distances between cell nuclei in the tissue increased. We could separate densely distributed nuclei in the hippocampus of mouse brain better compared to conventional UV-PAM. In short, we developed a novel PA imaging system with the enhanced temporal and spatial resolution by combining fast scanning modality and tissue expansion technology with UV-PAM.
In tumor resection surgery, the entire tumor must be removed to prevent local recurrence of cancer. To achieve effective and successful tumor resection surgery, an intraoperative examination is performed for quick decision-making during the surgical process. Examination of frozen sections is a common method, but it has limitations that it requires time-consuming tissue processing procedures which leads to interpretation errors. Photoacoustic microscopy (PAM) with ultraviolet (UV) laser is a promising intraoperative surgical margin assessment method that provides depth-resolved and label-free imaging of cell nuclei without sectioning and staining. Despite these advantages, conventional PAM still has limited imaging speed that does not allow real-time imaging, because it achieves the volumetric images by raster scanning using 2-axis step motors. To overcome the limitation, we developed a high-speed reflection-mode OR-PAM based on a UV scanner. Using the scanner module, it took 180 seconds to acquire one volumetric data over 1 × 1 mm2. In an in-vitro test, the measured lateral and axial resolution were 1.2 μm and 65.1 μm, respectively. We performed ex-vivo experiments on paraffin sections of tissues after deparaffinization that had been excised from a kidney, liver, colon-cancer and a liver-cancer patient. We could find structures in tumorous tissues distinguishable from normal tissues in 4 × 8 mm2 which is clinically meaningful FOV. We could also identify single nucleus in UV-PAM images, and match it with the corresponding nucleus in microscopic images.
Photoacoustic microscopy (PAM), an emerging biomedical imaging technology, has demonstrated the label-free imaging capability to visualize biomolecules with the aid of superior optical contrast in them. Especially employing ultraviolet (UV) laser at the wavelength of 266 nm, we have developed an UV-PAM. Unlike conventional histology methods such as frozen and formalin-fixed paraffin-embedded (FFPE) sections, UV-PAM can illustrate cell nuclei by utilizing superior light absorption of DNA/RNA without time consuming procedures. In-vitro experiments were conducted to evaluate the spatial resolutions of the developed system. The measured lateral resolution was 1.3 μm, and axial resolution was 62.2 μm. Then we performed ex-vivo experiments using frozen sections of mouse brain to demonstrate the imaging capability of UVPAM as a rapid histology tool. Oxidative stress induced by kainic acid (KA) was monitored using UV-PAM, which is considered as a significant cause for epileptic neuronal brain damage. We have shown the apoptotic feature resulted from the KA-induced hippocampal cell death in a mouse brain section. In contrast to the brain section of the control mouse model, the substantial nuclear marginalization of hippocampal cell death was illustrated in the vulnerable neurons of the CA1 and CA3 regions on the KA-treated mouse with PA imaging. In addition, the PA histologic results were evidenced by the corresponding HE stained images on both the control and the KA-treated mouse, showing similar hippocampal cell death. The PA histologic results could also provide its potential application for use in the monitoring of the morphological changes observed in astrocytes including hypotrophy, hyperplasia, and neoplasia. Further, it might be a beneficial histologic tool for treatment monitoring of neurodegenerative diseases such as acute traumatic brain injury and neuroprotective effects of treatments on the diseases.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.