中国计量科学研究院医学计量中心，北京 100029
目前，关于荧光流式细胞仪的计量和溯源方面的资料相对有限。为了研究基于荧光检测的流式细胞仪的技术发展并加快其标准化进程，对国内外文献资料进行整合，并总结了荧光流式细胞仪的类别、应用、相关标准化研究进展和关键参数的详细信息，主要包括仪器分辨率、散射光和荧光灵敏度、荧光线性相关系数、检出限、准确性、可重复性和稳定性等。国内外对广泛使用的几种评价荧光流式细胞仪性能的计量方法具有基本一致的标准。各个研究组织与应用领域对于流式细胞仪的性能具有不同的需求，一套完整可溯源的表征流式细胞术的计量标准及其评价方法能够方便实验室之间进行可重复和可比较的研究交流与讨论。
荧光检测 流式细胞仪 标准化 计量 溯源 激光与光电子学进展
2023, 60(4): 0400002
Author Affiliations
Abstract
Department of Electrical and Computer Engineering, University of Toronto, Toronto, ON, M5S 3G4, Canada
The Microchip Imaging Cytometer (MIC) is a class of integrated point-of-care detection systems based on the combination of optical microscopy and flow cytometry. MIC devices have the attributes of portability, cost-effectiveness, and adaptability while providing quantitative measurements to meet the needs of laboratory testing in a variety of healthcare settings. Based on the use of microfluidic chips, MIC requires less sample and can complete sample preparation automatically. Therefore, they can provide quantitative testing results simply using a finger prick specimen. The decreased reagent consumption and reduced form factor also help improve the accessibility and affordability of healthcare services in remote and resource-limited settings. In this article, we review recent developments of the Microchip Imaging Cytometer from the following aspects: clinical applications, microfluidic chip integration, imaging optics, and image acquisition. Following, we provide an outlook of the field and remark on promising technologies that may enable significant progress in the near future.The Microchip Imaging Cytometer (MIC) is a class of integrated point-of-care detection systems based on the combination of optical microscopy and flow cytometry. MIC devices have the attributes of portability, cost-effectiveness, and adaptability while providing quantitative measurements to meet the needs of laboratory testing in a variety of healthcare settings. Based on the use of microfluidic chips, MIC requires less sample and can complete sample preparation automatically. Therefore, they can provide quantitative testing results simply using a finger prick specimen. The decreased reagent consumption and reduced form factor also help improve the accessibility and affordability of healthcare services in remote and resource-limited settings. In this article, we review recent developments of the Microchip Imaging Cytometer from the following aspects: clinical applications, microfluidic chip integration, imaging optics, and image acquisition. Following, we provide an outlook of the field and remark on promising technologies that may enable significant progress in the near future.
microchip microfluidics flow cytometer imaging cytometer biosensors point-of-care testing biomedical engineering Opto-Electronic Advances
2022, 5(11): 210130
1 西安工业大学电子信息工程学院, 陕西 西安 710032
2 上海电力学院电子与信息工程学院, 上海 200090
3 中国工程物理研究院流体物理研究所, 四川 绵阳 621900
4 江南大学理学院光电信息科学与工程系, 江苏 无锡 214122
生物细胞的成像是生物和物理领域中的重要研究方向,尤其是大量细胞的测量与分析对于生物研究、疾病诊断具有重要的研究价值和意义。传统的显微检测仅局限于细胞形状的定性观察,且受视场限制,观察的细胞数量难以达到统计要求。将定量干涉显微和视场扫描结合,设计并实现了用于大量细胞成像与分析的定量干涉显微流式细胞仪。定量干涉显微可以恢复细胞的相位分布,结合视场扫描可以对大量细胞信息进行获取与分析。分别选用扩展主程序分析算法、正则化光学流场算法和傅里叶相位恢复算法,并结合不同的扫描方式,实现了对大量细胞相位面积、相位体积和圆率等不同参数的测量与统计。该系统有望在大通量高速细胞检测中获得应用。
全息 定量干涉显微 流式细胞仪 定量相位分布 细胞参数 激光与光电子学进展
2018, 55(12): 120901
Author Affiliations
Abstract
1 State Key Laboratory of Oncogenes and Related Genes Shanghai Cancer Institute Med-X Research Institute and School of Biomedical Engineering Shanghai Jiao Tong University 1954 Huashan Road, Shanghai 200030, P. R. China
2 Department of Biomedical Engineering Washington University in St. Louis One Brookings Drive, St. Louis, Missouri 63130, USA
3 Radiation Oncology Center Fudan University Shanghai Cancer Center (FUSCC) Shanghai 200032, P. R. China
Recent studies in oncology have addressed the importance of detecting circulating tumor cell clusters because circulating tumor cell clusters might survive and metastasize more easily than single circulating tumor cells. Signals with larger peak widths detected by in vivo flow cytometer (IVFC) have been used to identify cell clusters in previous studies. However, the accuracy of this criterion might be greatly degraded by variance in blood flow and the rolling behaviors of circulating tumor cells. Here, we propose a criterion and algorithm to distinguish cell clusters from single cells. In this work, we first used area-based and volume-based models for single fluorescent cells. Simulating each model, we analyzed the corresponding morphology of IVFC signals from cell clusters. According to the Rayleigh criterion, the valley between two adjacent peak signals from two distinguishable cells should be lower than 73.5% of the peak values. A novel signal processing algorithm for IVFC was developed based on this criterion. The results showed that cell clusters can be reliably identified using our proposed algorithm. Intravital imaging was also performed to further support our algorithm. With enhanced accuracy, IVFC is a powerful tool to study circulating cell clusters.
In vivo flow cytometer circulating tumor cell cell clusters signal processing algorithm computer simulation Rayleigh criterion Journal of Innovative Optical Health Sciences
2018, 11(5): 1850024
1 中国科学院长春光学精密机械与物理研究所, 吉林 长春 130033
2 长春长光思博光谱技术有限公司, 吉林 长春 130033
为了解决成像流式细胞仪多色、宽波段、长工作距离以及大数值孔径等难点，设计了1款多光谱成像流式细胞仪光学系统。系统基于模块化设计思想，灵活运用部分和整体的优化方法，保证各模块的相对独立性；显微物镜引入衍射元件对宽谱段色差进行校正，保证在全波段下成像质量均达到理想的性能指标；多光谱分解镜组采用二向色镜堆栈分光，多重结构同时优化6个通道，大大降低了系统对色差的校正难度。全局优化得到最终的光学系统，放大倍率为60倍，视场为60 μm×128 μm，波段为420~800 nm，6个分光通道，分辨率达到0.5 μm，成像质量接近衍射极限。
光学设计 成像流式细胞仪 宽波段
1 合肥工业大学 仪器科学与光电工程学院, 安徽 合肥 230009
2 北京信息科技大学 光电测试技术北京市重点实验室, 北京 100101
提出了一种基于90° Mie散射的高速图像采集微球测速方法, 用于准确评估流式细胞仪流动室内的层流状态及单细胞流的稳定性。利用流动室内微球速度的稳定性对流动室内单细胞流的稳定性进行了评估。首先, 利用高速显微成像系统采集90° Mie散射光的图像, 选取90°侧向散射光以避免激发光源直射光的干扰, 同时去除背景光源并提高图像对比度; 然后, 利用基于梯形白化权函数的灰色聚类分析方法对微球拖尾图像进行分类, 实现对不足、正常、衍射和重叠4种情况的准确分类; 最后, 利用中点法确定正常图像上升沿及下降沿的边界, 提高拖尾长度计算的准确性。搭建了高速微球测速实验系统, 对本文方法进行验证。结果表明, 该方法能够获得清晰的微球拖尾图像并对微球拖尾图像进行准确分类。对本文实验系统测得的微球拖尾长度平均值为116.9个像素点, 标准差为1.7。
流式细胞仪 液路稳定性评估 微球测速 高速图像采集 聚类分析 flow cytometer flow stability evaluation particle velocity measurement high-speed image sampling cluster analysis
Author Affiliations
Abstract
1 Department of Chemistry, Fudan University, Shanghai, P. R. China
2 Institutes of Biomedical Sciences, Fudan University, Shanghai, P. R. China
3 Department of Urology, Xinhua Hospital Shanghai Jiao Tong University 1665, Kongjiang Road, Shanghai 200092, P. R. China
4 Med-X Research Institute, Shanghai Jiao Tong University 1954, Huashan Road, Shanghai 200240, P. R. China
5 School of Biomedical Engineering Shanghai Jiao Tong University, Shanghai, P. R. China
Metastasis is a very complicated multi-step process and accounts for the low survival rate of the cancerous patients. To metastasize, the malignant cells must detach from the primary tumor and migrate to secondary sites in the body through either blood or lymph circulation. Macrophages appear to be directly involved in tumor progression and metastasis. However, the role of macrophages in affecting cancer metastasis has not been fully elucidated. Here, we have utilized an emerging technique, namely in vivo flow cytometry (IVFC) to study the depletion kinetics of circulating prostate cancer cells in mice and determine how depletion of macrophages by the liposome-encapsulated clodronate affects the depletion kinetics. Our results show different depletion kinetics of PC-3 cells between the macrophage-deficient group and the control group. The number of circulating tumor cells (CTCs) in the macrophage-deficient group decreases in a slower manner compared to the control mice group. The differences in depletion kinetics indicate that the absence of macrophages facilitates the stay of prostate cancer cells in circulation. In addition, our imaging data suggest that macrophages might be able to arrest, phagocytose and digest PC-3 cells. Therefore, phagocytosis may mainly contribute to the depletion kinetic differences. The developed methods elaborated here would be useful to study the relationship between macrophages and tumor metastasis in small animal cancer models.
Prostate cancer macrophages liposome-encapsulated clodronate in vivo flow cytometer circulating tumor cells Journal of Innovative Optical Health Sciences
2012, 5(4): 1250027
