Tel Aviv University, Department of Biomedical Engineering, Tel Aviv, Israel.
University of Münster, Biomedical Technology Center, Münster, Germany.
J Biomed Opt. 2017 Jun 1;22(6):66012. doi: 10.1117/1.JBO.22.6.066012.
We present highly dynamic photothermal interferometric phase microscopy for quantitative, selective contrast imaging of live cells during flow. Gold nanoparticles can be biofunctionalized to bind to specific cells, and stimulated for local temperature increase due to plasmon resonance, causing a rapid change of the optical phase. These phase changes can be recorded by interferometric phase microscopy and analyzed to form an image of the binding sites of the nanoparticles in the cells, gaining molecular specificity. Since the nanoparticle excitation frequency might overlap with the sample dynamics frequencies, photothermal phase imaging was performed on stationary or slowly dynamic samples. Furthermore, the computational analysis of the photothermal signals is time consuming. This makes photothermal imaging unsuitable for applications requiring dynamic imaging or real-time analysis, such as analyzing and sorting cells during fast flow. To overcome these drawbacks, we utilized an external interferometric module and developed new algorithms, based on discrete Fourier transform variants, enabling fast analysis of photothermal signals in highly dynamic live cells. Due to the self-interference module, the cells are imaged with and without excitation in video-rate, effectively increasing signal-to-noise ratio. Our approach holds potential for using photothermal cell imaging and depletion in flow cytometry.
我们提出了一种高度动态的光热干涉相位显微镜，用于在流动过程中对活细胞进行定量、选择性对比成像。金纳米粒子可以被生物功能化以与特定的细胞结合，并由于等离子体共振而被刺激产生局部温度升高，导致光学相位的快速变化。这些相位变化可以通过干涉相位显微镜记录下来，并进行分析，以形成细胞中纳米粒子结合位点的图像，从而获得分子特异性。由于纳米粒子的激发频率可能与样品动力学频率重叠，因此在静止或缓慢动态的样品上进行光热相位成像。此外，光热信号的计算分析非常耗时。这使得光热成像不适合需要动态成像或实时分析的应用，例如在快速流动过程中分析和分选细胞。为了克服这些缺点，我们利用外部干涉模块并开发了新的算法，基于离散傅里叶变换变体，能够快速分析高度动态活细胞中的光热信号。由于自干涉模块，细胞在视频速率下进行激发和不激发成像，有效地提高了信噪比。我们的方法在流动细胞术中用光热细胞成像和耗尽方面具有潜力。
