Division of Neurosurgery, Department of Surgery, Neuroscience Institute, University of Cape Town, Cape Town, South Africa.
Department of Internal Medicine and Radboud Center of Infectious Diseases, Radboud University Medical Center, Nijmegen, The Netherlands.
Sci Rep. 2024 Jan 30;14(1):2463. doi: 10.1038/s41598-024-52669-1.
The pauci-cellular nature of cerebrospinal (CSF), particularly ventricular CSF, and the rapid cell death following sampling, incumbers the use of flow cytometric analysis of these samples in the investigation of central nervous system (CNS) pathologies. Developing a method that allows long-term storage and batched analysis of CSF samples without compromising cell integrity is highly desirable in clinical research, given that CSF is often sampled after hours creating logistical difficulties for fresh processing. We examined percentages and relative proportion of peripheral and brain-derived immune cells in cryopreserved and transfix-treated CSF, compared to freshly processed CSF. Cell proportions were more comparable between Fresh and Cryopreserved CSF (mean of differences = 3.19), than between fresh and transfix-treated CSF (mean of differences = 14.82). No significant differences in cell percentages were observed in fresh versus cryopreserved CSF; however significantly lower cell percentages were observed in transfix-treated CSF compared to Fresh CSF [(CD11b (p = 0.01), CD4 (p = 0.001), CD8 (p = 0.007), NK cells (p = 0.04), as well as CD69 activation marker (p = 0.001)]. Furthermore, loss of marker expression of various lymphocyte sub-populations were observed in transfix-treated CSF. Cryopreservation is a feasible option for long-term storage of ventricular CSF and allows accurate immunophenotyping of peripheral and brain-derived cell populations by flow cytometry.
脑脊髓液（CSF），尤其是脑室 CSF 的细胞数量较少，且采样后细胞迅速死亡，这给利用流式细胞术分析这些样本以研究中枢神经系统（CNS）病变带来了困难。开发一种能够长期储存 CSF 样本并进行批量分析而不影响细胞完整性的方法，在临床研究中是非常需要的，因为 CSF 通常在非工作时间采集，这给及时处理带来了后勤上的困难。我们比较了冷冻保存和 TransFix 处理 CSF 与新鲜处理 CSF 中外周和脑源性免疫细胞的百分比和相对比例。与新鲜 CSF 相比，冷冻保存 CSF 中细胞比例更接近（差异平均值为 3.19），而新鲜 CSF 与 TransFix 处理 CSF 之间的差异平均值为 14.82。新鲜 CSF 与冷冻保存 CSF 之间的细胞百分比无显著差异；然而，与新鲜 CSF 相比，TransFix 处理 CSF 中的细胞百分比显著降低（CD11b（p=0.01）、CD4（p=0.001）、CD8（p=0.007）、NK 细胞（p=0.04）以及 CD69 激活标志物（p=0.001））。此外，TransFix 处理 CSF 中观察到各种淋巴细胞亚群的标记表达丢失。冷冻保存是长期储存脑室 CSF 的可行选择，并且允许通过流式细胞术对外周和脑源性细胞群进行准确的免疫表型分析。
