Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, and Chemical Biology Center, College of Chemistry, Central China Normal University, Wuhan, 430079, Hubei, China.
State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Institute of Physics and Mathematics, Chinese Academy of Sciences, Wuhan, 430071, Hubei, China.
Anal Bioanal Chem. 2019 May;411(13):2767-2780. doi: 10.1007/s00216-019-01738-w. Epub 2019 Apr 11.
Blood counting is one of the most commonly ordered clinical assays, and is often part of the basis for initial diagnosis and screening for disease. While substantial prior research has shown the ability of portable instruments to accurately produce blood counts through image- or flow-based cytometry, these methods require complex sample preparation using either costly commercial imaging chambers or complicated reagents. To address these issues, in this paper we developed a method to prepare trace volumes of whole blood aimed at portable blood counting. The strategy is based on pre-storing dry-form reagents and fabricating a specifically designed cell counter. In order to obtain total cell counts for red blood cells, platelets, and 3-part differentials of white blood cells, two parallel counting chambers with different depths are made from cost- and environmentally friendly materials using soft lithography. As little as 1 μl of whole blood is prepared with pre-stored reagents in centrifuge vials, whereas red blood cells are sphered and white blood cells are stained at the same time. Driven by the capillary force, prepared blood samples enter the hydrophilic chambers automatically. Monolayers of cells are formed when the blood dilution factors and the chamber depths are co-optimized. Combined with our previous custom-built instrument and automated analysis algorithm, the sample preparation strategy allows producing counting results with excellent agreement to a gold-standard clinical hematology instrument. The success of this preparation method may further advance applications of our technology for global use in low-resource settings where central hematology laboratories are not accessible. Graphical abstract Graphical Abstract.
血细胞计数是最常进行的临床检测之一，通常也是疾病初始诊断和筛查的基础。虽然已有大量研究表明，便携式仪器能够通过基于图像或流动的细胞计数准确地生成血细胞计数，但这些方法需要使用昂贵的商用成像室或复杂的试剂进行复杂的样本制备。为了解决这些问题，我们在本文中开发了一种用于便携式血细胞计数的微量全血制备方法。该策略基于预先储存干粉试剂和制造专门设计的细胞计数器。为了获得红细胞、血小板和白细胞三分群的总细胞计数，我们使用软光刻技术，用成本效益高且环保的材料制作了两个具有不同深度的平行计数室。只需在微量离心管中预先储存试剂即可准备 1 μl 的全血，同时实现红细胞的球化和白细胞的染色。在毛细作用力的驱动下，制备好的血样自动进入亲水室。当血液稀释因子和腔室深度协同优化时，会形成单层细胞。结合我们之前的定制仪器和自动化分析算法，该样本制备策略可以得到与金标准临床血液学仪器非常吻合的计数结果。这种制备方法的成功可能会进一步推动我们的技术在全球范围内的应用，特别是在难以获得中央血液学实验室的资源有限的环境中。
