Photothermal Imaging of Live Cells

生物组织的激光成像代表了前所未有的最新进展，它已经彻底改变了生物医学的许多领域，特别是细胞生物学和癌症诊断。然而，这些技术不适用于广泛的生物医学任务，需要可视化的非荧光，低散射，低折射吸收结构。该提案的最终目的是通过开发激光光热（PT）成像（PTI）克服现有技术的局限性，该成像具有高吸收灵敏度，可用于在其自然状态下对非荧光活细胞进行光谱成像，而无需标准染色和扫描，包括时间分辨监测全细胞的综合PT响应的能力。本论文的具体目标如下：目标1：研制一种新型的激光PT相衬显微镜/光谱仪，并对其灵敏度和空间分辨率进行评估。目的2：研究激光与PTI的细胞相互作用，确定其在活细胞中的非侵入性和侵入性参数。目的3：开发PT对比探针的PTI，例如金纳米颗粒。目的4：评价PT技术将常规荧光探针可视化为PT探针的能力。目的5：研究PT活细胞的光谱/显微镜能力。与现有工具相比，新PT技术的潜在优势包括在其天然状态下对非荧光细胞进行高灵敏度，高分辨率成像，而无需染色和扫描。这些技术将扩大我们对正常和病理状态下细胞微观结构特性的理解，并可应用于细胞癌症诊断，PT敏化剂选择性癌症治疗的指导，药物-细胞相互作用的研究，药物递送的控制和一些光疗干预。PTI的技术实现相对简单，需要结合传统的光学相衬显微镜和已经常规使用的纳秒激光。

DESCRIPTION (provided by applicant): Laser imaging of biotissue represents an unprecedented recent advance that has revolutionized many areas of biomedicine, especially cell biology and cancer diagnosis. Nevertheless, these techniques are not applicable to broad biomedical tasks requiring visualization of nonfluorescent, low-scattering, and low-refractive absorbing structures. The ultimate aim of this proposal is to overcome the limitations of existing technologies by developing laser photothermal (PT) imaging (PTI) that has high absorption sensitivity for spectral imaging of nonfluorescent living cells in their natural state without standard staining and scanning, including the capability for time-resolved monitoring of the integrated PT response from whole cells. The Specific Aims of this proposal are as follows: Aim 1: Develop a new modification of the laser PT phase-contrast microscope/spectrometer, and estimate its sensitivity and spatial resolution. Aim 2: Study laser-cell interactions with PTI to determine its noninvasive and invasive parameters in living cells. Aim 3: Develop PTI of PT contrast probes, such as gold nanoparticles. Aim 4: Evaluate the ability of the PT technique to visualize conventional fluorescent probes as PT probes. Aim 5: Investigate PT the capability of spectroscopy/microscopy of living cells. The potential advantages of a new PT technology, compared with existing tools, include high-sensitivity, high-resolution imaging of nonfluorescent cells in their native state, without the need for staining and scanning. These techniques will enlarge our understanding of cell microstructure properties in normal and pathologic states and can be applied to cellular cancer diagnostics, guidance of selective cancer therapy with a PT sensitizer, study of drug-cell interactions, control of drug delivery, and some phototherapeutic interventions. The technical realization of PTI is relatively simple, requiring the combination of conventional optical phase-contrast microscopy and the already routinely used nanosecond laser.