University of Arkansas, Department of Biomedical Engineering, Fayetteville, Arkansas, United States.
University of Arkansas, Pat Walker Health Center, Fayetteville, Arkansas, United States.
J Biomed Opt. 2017 Mar 1;22(3):35001. doi: 10.1117/1.JBO.22.3.035001.
There exists a broad range of techniques that can be used to classify and count white blood cells in a point-of-care (POC) three-part leukocyte differential test. Improvements in lenses, light sources, and cameras for image-based POC systems have renewed interest in acridine orange (AO) as a contrast agent, whereby subpopulations of leukocytes can be differentiated by colorimetric analysis of AO fluorescence emission. We evaluated the effect on test accuracy using different AO staining and postprocessing methods in the context of an image-based POC colorimetric cell classification scheme. Thirty blood specimens were measured for percent cell counts using our POC system and a conventional hematology analyzer for comparison. Controlling the AO concentration used during whole-blood staining, the incubation time with AO, and the colorimetric ratios among the three population of leukocytes yielded a percent deviation of 0.706%, ? 1.534 % , and ? 0.645 % for the lymphocytes, monocytes, and granulocytes, respectively. Overall, we demonstrated that a redshift in AO fluorescence was observed at elevated AO concentrations, which lead to reproducible inaccuracy of cell counts. This study demonstrates there is a need for a strict control of the AO staining and postprocessing methods to improve test accuracy in these POC systems.
存在广泛的技术可用于在即时护理（POC）三分群白细胞分类测试中对白细胞进行分类和计数。基于图像的 POC 系统中镜头、光源和相机的改进重新激发了人们对吖啶橙（AO）作为对比剂的兴趣，通过对 AO 荧光发射的比色分析可以区分白细胞的亚群。我们评估了在基于图像的 POC 比色细胞分类方案的背景下，使用不同的 AO 染色和后处理方法对测试准确性的影响。使用我们的 POC 系统和传统血液分析仪测量了 30 个血液样本的细胞百分比计数，以进行比较。通过控制全血染色过程中 AO 的浓度、与 AO 的孵育时间以及三种白细胞群之间的比色比值，淋巴细胞、单核细胞和粒细胞的百分比偏差分别为 0.706%、±1.534%和±0.645%。总的来说，我们证明在 AO 浓度升高时观察到 AO 荧光的红移，这导致细胞计数的重复性不准确。这项研究表明，需要严格控制 AO 染色和后处理方法，以提高这些 POC 系统中的测试准确性。
