Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, United States.
Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, United States.
Talanta. 2019 Nov 1;204:731-738. doi: 10.1016/j.talanta.2019.05.096. Epub 2019 Jun 1.
Blood is a routinely tested biological fluid for diagnosis and monitoring of diseases as many diseases would trigger a change in white blood cell count. Thus, several methods have been established to isolate or enrich white blood cells from patient blood samples for such analyses. One method of preparing an enriched white blood cell sample is through the selective lysis of red blood cells by hypotonic shock and restoration of osmolarity to maintain viability of target white blood cells. An inherent problem with this approach is the loss of target cells during sample handling. We report a two-stage separation system that can perform lysis and restoration of osmolarity of blood on-chip and direct the resultant sample to the second step of the analysis. Hence, there is no loss of sample. The post-lysis makeup features a protein-rich buffer to help stabilize cells. As proof of concept, we spiked HL-60 cells into a whole blood and a pre-lysed blood sample and compared capture metrics of each method using a downstream affinity separation. The capture efficiency of the whole blood sample ranged between 40 and 80% using <7 μL of sample compared to 10-52% from 60 μL of pre-lysed blood required for similar analysis. In addition, both pre-lysed and whole blood samples showed no significant difference in purity and viability. This two-stage separation system has demonstrated the capacity to replace centrifugation and wash steps required for the preparation of lysed blood, for white blood cell analyses.
血液是一种常用于诊断和监测疾病的常规检测生物样本，因为许多疾病都会导致白细胞计数发生变化。因此，已经建立了几种方法来从患者的血液样本中分离或富集白细胞，以进行此类分析。一种制备富含白细胞样本的方法是通过低渗休克选择性裂解红细胞，并恢复渗透压以维持靶白细胞的活力。这种方法的一个固有问题是在样本处理过程中靶细胞的丢失。我们报告了一种两阶段分离系统，该系统可以在芯片上进行溶血和渗透压恢复，并将得到的样本直接导向分析的第二步。因此，没有样本损失。溶血后的组成部分具有富含蛋白质的缓冲液，有助于稳定细胞。作为概念验证，我们将 HL-60 细胞掺入全血和预溶血的血液样本中，并使用下游亲和分离比较每种方法的捕获指标。与使用 60 μL 预溶血血液进行类似分析所需的 10-52%相比，使用<7 μL 的样本，全血样本的捕获效率在 40%到 80%之间。此外，预溶血和全血样本在纯度和活力方面均无显著差异。这种两阶段分离系统已经证明了能够替代为制备溶血血液而进行的离心和洗涤步骤，用于白细胞分析。
