细胞外囊泡(EV)被认为在神经变性的进展中发挥作用,并且能够将病理蛋白从一个细胞传递到另一个细胞。可以分离EV的生物流体之一是脑脊液(CSF)。然而,到目前为止,对少量CSF进行的研究很少.由于合并患者样本可能会导致患者个人基本信息的丢失,脑脊液样本很珍贵,重要的是要有有效的技术来将电动汽车从较小的体积中分离出来。在这项研究中,已为此目的评估了SystemBiosciences的SmartSECHT分离试剂盒。SmartSECHT分离试剂盒用于从500μL初始体积的CSF中分离EV,导致两个可能的EV分数为500μL。使用整个范围的表征技术对两种级分进行表征并相互比较。结果表明,这两个部分都存在电动汽车,尽管分数1显示出更多的重现性结果超过不同的表征方法。例如,CMG(CellMask绿色膜染色)荧光纳米裂纹分析(NTA),ExoView,并且颗粒/μg比率表明级分1和2之间存在明显差异,其中级分1以污染最少的方式洗脱出大多数EV。在其他方法中,这种差异不太明显。我们仅使用500μL的CSF起始体积成功地进行了互补表征测试,and,得出的结论是,部分1由足够纯的电动汽车组成,用于进一步的生物标志物研究。这意味着未来的EV提取可能基于较小的CSF量,例如来自个体患者。这样,患者样本不必合并,患者的个体信息可纳入即将进行的研究,可能链接EV内容,个性化神经系统诊断的大小和分布。
Extracellular vesicles (EVs) are suggested to have a role in the progression of neurodegeneration, and are able to transmit pathological proteins from one cell to another. One of the biofluids from which EVs can be isolated is cerebrospinal fluid (CSF). However, so far, few studies have been performed on small volumes of CSF. Since pooling of patient samples possibly leads to the loss of essential individual patient information, and CSF samples are precious, it is important to have efficient techniques for the isolation of EVs from smaller volumes. In this study, the SmartSEC HT isolation kit from System Biosciences has been evaluated for this purpose. The SmartSEC HT isolation kit was used for isolation of EVs from 500 μL starting volumes of CSF, resulting in two possible EV fractions of 500 μL. Both fractions were characterised and compared to one another using a whole range of characterisation techniques. Results indicated the presence of EVs in both fractions, albeit fraction 1 showed more reproducible results over the different characterisation methods. For example, CMG (CellMask Green membrane stain) fluorescence nanotracking analysis (NTA), ExoView, and the particles/μg ratio demonstrated a clear difference between fraction 1 and 2, where fraction 1 came out as the one where most EVs were eluted with the least contamination. In the other methods, this difference was less noticeable. We successfully performed complementary characterisation tests using only 500 μL of CSF starting volume, and, conclude that fraction 1 consisted of sufficiently pure EVs for further biomarker studies. This means that future EV extractions may be based upon smaller CSF quantities, such as from individual patients. In that way, patient samples do not have to be pooled and individual patient information can be included in forthcoming studies, potentially linking EV content, size and distribution to individualised neurological diagnoses.