Ymeti Aurel, Li Xiao, Lunter Björn, Breukers Christian, Tibbe Arjan G J, Terstappen Leon W M M, Greve Jan
Biophysical Engineering Group, MESA+ Institute for Nanotechnology and Biomedical Technology Institute, Faculty of Science and Technology, University of Twente, 7500 AE Enschede, The Netherlands.
Cytometry A. 2007 Mar;71(3):132-42. doi: 10.1002/cyto.a.20375.
For resource-poor countries, affordable methods are required for enumeration of CD4(+) T lymphocytes of HIV-positive patients. For infants, additional determination of CD4/CD8 ratio is needed.
We determine the CD4(+) and CD8(+) T lymphocytes as the CD3(+)CD4(+) and CD3(+)CD8(+) population of blood cells. Target cells are CD3-immunomagnetically separated from the whole blood, and CD4-Phycoerythrin and CD8-PerCP immunofluorescently labeled. A point-of-care single platform image cytometer was developed to enumerate the target CD3(+)CD4(+) and CD3(+)CD8(+) populations. It has light-emitting diodes illumination, is fully computer-controlled, operates from a 12 V battery, and was designed to be cheap and easy-to-handle. Target cells are imaged on a CCD camera and enumerated by an image analysis algorithm. The cytometer outputs the absolute number of CD4(+) and CD8(+) T lymphocytes/microl and CD4/CD8 ratio.
The quality of the cell images obtained with the cytometer is sufficient for a reliable enumeration of target cells. The image cytometer achieves an accuracy of better than 10% in the range of 50-1700 cells/microl. Analysis of blood samples from HIV patients yields a good agreement with the TruCount method for CD4 and CD8 count and CD4/CD8 ratio.
The image cytometer is affordable (component costs $3,000), compact (25 x 25 x 20 cm(3)), and uses disposable test materials, making it a good candidate to monitor progression of immunodeficiency disease in resource-poor settings.
对于资源匮乏的国家而言，需要有经济实惠的方法来对HIV阳性患者的CD4(+) T淋巴细胞进行计数。对于婴儿，则需要额外测定CD4/CD8比值。
我们将CD4(+)和CD8(+) T淋巴细胞确定为血细胞中的CD3(+)CD4(+)和CD3(+)CD8(+)群体。通过CD3免疫磁珠法从全血中分离出靶细胞，并用CD4-藻红蛋白和CD8-藻红蛋白-氰化物荧光素进行标记。开发了一种即时检测单平台图像细胞仪来对靶细胞CD3(+)CD4(+)和CD3(+)CD8(+)群体进行计数。它采用发光二极管照明，完全由计算机控制，使用12伏电池供电，设计得既便宜又易于操作。靶细胞在电荷耦合器件相机上成像，并通过图像分析算法进行计数。该细胞仪输出CD4(+)和CD8(+) T淋巴细胞每微升的绝对数量以及CD4/CD8比值。
用该细胞仪获得的细胞图像质量足以可靠地计数靶细胞。该图像细胞仪在50 - 1700个细胞/微升的范围内，准确度优于10%。对HIV患者血样的分析结果与TruCount方法在CD4和CD8计数以及CD4/CD8比值方面具有良好的一致性。
该图像细胞仪价格实惠（组件成本为3000美元），体积小巧（25×25×20立方厘米），且使用一次性检测材料,使其成为在资源匮乏地区监测免疫缺陷疾病进展的理想选择。
