Department of Biomedical Engineering, Southern University of Science and Technology, Shenzhen 518055, China.
Shenzhen Bay Laboratory, Shenzhen 518132, China.
ACS Nano. 2022 Feb 22;16(2):3231-3238. doi: 10.1021/acsnano.1c11103. Epub 2022 Jan 26.
Photoacoustic imaging (PAI) has been widely used in multiscale and multicontrast imaging of biological structures and functions. Optical resolution photoacoustic microscopy (OR-PAM), an emerging submodality of PAI, features high lateral resolution and rich optical contrast, indicating great potential in visualizing cellular and subcellular structures. However, three-dimensional (3D) imaging of subcellular structures using OR-PAM has remained a challenge due to the limited axial resolution. In this study, we propose a multicolor 3D photoacoustic microscopy with high lateral/axial resolutions of 0.42/2 and 0.5/2.5 μm at 532 and 780 nm excitation, respectively. Owing to the significantly increased axial resolution, we could visualize the volumetric subcellular structures of melanoma cells using intrinsic contrast. In addition, we carried out multicolor imaging of labeled microtubules/clathrin-coated pits (CCP) and microtubules/mitochondria, respectively, with one scanning by using two different excitation wavelengths. The internal connections between different subcellular structures are revealed by quantitatively comparing the spatial distributions of microtubules/CCP and microtubules/mitochondria in a single cell. Current results suggest that the proposed OR-PAM may serve as an efficient tool for subcellular and cytophysiological studies.
光声成像是一种广泛应用于生物结构和功能的多尺度和多对比度成像技术。光分辨光声显微镜（OR-PAM）是光声成像的一个新兴子领域，具有高横向分辨率和丰富的光学对比度，在可视化细胞和亚细胞结构方面具有巨大的潜力。然而，由于轴向分辨率有限，使用 OR-PAM 进行亚细胞结构的三维成像仍然是一个挑战。在这项研究中，我们提出了一种多色三维光声显微镜，在 532nm 和 780nm 激发下，具有 0.42/2μm 和 0.5/2.5μm 的高横向/轴向分辨率。由于轴向分辨率显著提高，我们可以使用固有对比度可视化黑色素瘤细胞的体积亚细胞结构。此外，我们分别使用两个不同的激发波长进行了标记微管/网格蛋白包被凹陷（CCP）和微管/线粒体的多色成像，仅通过一次扫描即可实现。通过定量比较单个细胞中微管/CCP 和微管/线粒体的空间分布，揭示了不同亚细胞结构之间的内部联系。目前的结果表明，所提出的 OR-PAM 可能成为亚细胞和细胞生理学研究的有效工具。
