Lourbopoulos Athanasios, Müller Stephan A, Jocher Georg, Wick Manfred, Plesnila Nikolaus, Lichtenthaler Stefan F
Laboratory of Experimental Stroke Research, Institute for Stroke and Dementia Research (ISD), LMU University Hospital, Munich, Germany.
German Center for Neurodegenerative Diseases (DZNE), Munich, Germany; Neuroproteomics, School of Medicine and Health, Klinikum rechts der Isar, Technical University of Munich, Munich, Germany.
Mol Cell Proteomics. 2025 Mar 27;24(5):100958. doi: 10.1016/j.mcpro.2025.100958.
The mouse is the species most commonly used in preclinical research, but protein analytics of murine cerebrospinal fluid (CSF) remains challenging because of low sample volumes (often <10 μl) and frequent contaminations with blood. We developed an improved CSF sampling method that allows routine collection of larger volumes (20-30 μl) of pure CSF from individual mice, enabling multiple protein analytical assays from a single sample. Based on cell counts and hemoglobin ELISAs, we provide an easy quality control workflow for obtaining cell- and blood-free murine CSF. Through mass spectrometry-based proteomics using an absolutely quantified external standard, we estimated concentrations for hundreds of mouse CSF proteins. While repeated CSF sampling from the same mouse was possible, it induced CSF proteome changes. Applying the improved method, we found that the mouse CSF proteome remains largely stable over time in wild-type mice, but that amyloid pathology in the 5xFAD mouse model of Alzheimer's disease massively changes the CSF proteome. Neurofilament light chain and TREM2, markers of neurodegeneration and activated microglia, respectively, were strongly upregulated and validated using immunoassays. In conclusion, our refined murine CSF collection method overcomes previous limitations, allowing multiple quantitative protein analyses for applications in biomedicine.
小鼠是临床前研究中最常用的物种，但由于样本量低（通常<10μl）且经常被血液污染，对小鼠脑脊液（CSF）进行蛋白质分析仍然具有挑战性。我们开发了一种改进的脑脊液采样方法，该方法可以从单个小鼠中常规收集更大体积（20 - 30μl）的纯脑脊液，从而能够对单个样本进行多种蛋白质分析测定。基于细胞计数和血红蛋白酶联免疫吸附测定，我们提供了一种简单的质量控制工作流程，以获得无细胞和无血液的小鼠脑脊液。通过使用绝对定量外标物的基于质谱的蛋白质组学，我们估计了数百种小鼠脑脊液蛋白质的浓度。虽然可以对同一只小鼠重复进行脑脊液采样，但这会引起脑脊液蛋白质组的变化。应用改进后的方法，我们发现野生型小鼠的脑脊液蛋白质组随时间推移在很大程度上保持稳定，但阿尔茨海默病5xFAD小鼠模型中的淀粉样病理改变会极大地改变脑脊液蛋白质组。神经丝轻链和TREM2分别是神经退行性变和活化小胶质细胞的标志物，它们均显著上调，并通过免疫测定进行了验证。总之，我们改进的小鼠脑脊液收集方法克服了以前的局限性，允许进行多种定量蛋白质分析，以应用于生物医学领域。
