Georgia Institute of Technology, School of Electrical and Computer Engineering, Atlanta, Georgia, United States.
Georgia Institute of Technology and Emory University, Wallace H. Coulter Department of Biomedical En, United States.
J Biomed Opt. 2022 Jun;27(6). doi: 10.1117/1.JBO.27.6.066502.
Quantitative oblique back-illumination microscopy (qOBM) is a recently developed label-free imaging technique that enables 3D quantitative phase imaging of thick scattering samples with epi-illumination. Here, we propose dynamic qOBM to achieve functional imaging based on subcellular dynamics, potentially indicative of metabolic activity. We show the potential utility of this novel technique by imaging adherent mesenchymal stromal cells (MSCs) grown in bioreactors, which can help address important unmet needs in cell manufacturing for therapeutics.
We aim to develop dynamic qOBM and demonstrate its potential for functional imaging based on cellular and subcellular dynamics.
To obtain functional images with dynamic qOBM, a sample is imaged over a period of time and its temporal signals are analyzed. The dynamic signals display an exponential frequency response that can be analyzed with phasor analysis. Functional images of the dynamic signatures are obtained by mapping the frequency dynamic response to phasor space and color-coding clustered signals.
Functional imaging with dynamic qOBM provides unique information related to subcellular activity. The functional qOBM images of MSCs not only improve conspicuity of cells in complex environments (e.g., porous micro-carriers) but also reveal two distinct cell populations with different dynamic behavior.
In this work we present a label-free, fast, and scalable functional imaging approach to study and intuitively display cellular and subcellular dynamics. We further show the potential utility of this novel technique to help monitor adherent MSCs grown in bioreactors, which can help achieve quality-by-design of cell products, a significant unmet need in the field of cell therapeutics. This approach also has great potential for dynamic studies of other thick samples, such as organoids.
定量斜向背照明显微镜（qOBM）是一种最近开发的无标记成像技术，可通过 epi 照明对厚散射样品进行 3D 定量相位成像。在这里，我们提出动态 qOBM，以基于亚细胞动力学实现功能成像，这可能表明代谢活性。我们通过对在生物反应器中生长的贴壁间充质基质细胞（MSCs）进行成像来展示这种新技术的潜在应用，这有助于解决细胞治疗中制造方面的一些重要未满足需求。
我们旨在开发动态 qOBM，并证明其基于细胞和亚细胞动力学进行功能成像的潜力。
为了用动态 qOBM 获得功能图像，对样本进行一段时间的成像，并对其时间信号进行分析。动态信号显示出指数频率响应，可以通过相向量分析进行分析。通过将频率动态响应映射到相空间并对聚类信号进行颜色编码，可以获得动态特征的功能图像。
动态 qOBM 的功能成像提供了与亚细胞活性相关的独特信息。MSCs 的功能 qOBM 图像不仅提高了复杂环境（例如多孔微载体）中细胞的对比度，而且还揭示了具有不同动态行为的两种不同细胞群体。
在这项工作中，我们提出了一种无标记、快速且可扩展的功能成像方法，用于研究和直观显示细胞和亚细胞动力学。我们进一步展示了这项新技术的潜在应用，以帮助监测在生物反应器中生长的贴壁 MSCs，这有助于实现细胞产品的质量设计，这是细胞治疗领域的一个重大未满足需求。这种方法对于其他厚样品（如类器官）的动态研究也具有很大的潜力。
