Lipidomics focuses on the comprehensive analysis of lipids and their interactions with other molecules. Many biological samples are available in small volumes or limited amounts and display very complex lipid compositions; thus, highly sensitive lipidomic analysis methods are often needed. NanoLC-MS offers extremely high sensitivity, although it is known to be technically more challenging than the conventional LC-MS approach, requiring greater care and maintenance. This work describes the development and optimization of a nanoLC-MS method for routine analysis of the lipidome of small volumes of biological samples. We focused on achieving robust conditions for high sensitivity analysis of complex samples. The nanoLC method, mass spectrometry conditions and sample preparation by liquid-liquid extraction of lipids were fully optimized using serum samples and deuterated lipid standards, including an evaluation of contamination sources. The performance of the method was assessed through the analysis of human and pig sera, as well as cerebrospinal fluid samples from pigs. This method allowed the detection of 9900 to 12,200 features by employing only 1.0-2.5 μL of serum samples and identification of 5842 lipids from 36 subclasses within a 50-min gradient. The method can be easily adapted to other types of biological samples where only limited volumes are available.

中文翻译：

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用于高灵敏度全球脂质组学分析的 NanoLC-MS 工作流程的开发

脂质组学侧重于综合分析脂质及其与其他分子的相互作用。许多生物样品体积小或数量有限，显示出非常复杂的脂质成分；因此，通常需要高度灵敏的脂质组学分析方法。NanoLC-MS 具有极高的灵敏度，但众所周知，它在技术上比传统的 LC-MS 方法更具挑战性，需要更多的维护和保养。这项工作描述了用于对小体积生物样品的脂质组进行常规分析的 nanoLC-MS 方法的开发和优化。我们专注于实现复杂样品的高灵敏度分析的稳健条件。纳米液相色谱法，使用血清样品和氘代脂质标准品对质谱条件和脂质液-液萃取样品制备进行了全面优化，包括对污染源的评估。通过分析人和猪的血清以及猪的脑脊液样品来评估该方法的性能。该方法仅使用 1.0-2.5 μL 的血清样本即可检测 9900 到 12,200 个特征，并在 50 分钟的梯度内鉴定来自 36 个亚类的 5842 种脂质。该方法可以很容易地适用于其他类型的生物样品，其中只有有限的体积可用。以及猪的脑脊液样本。该方法仅使用 1.0-2.5 μL 的血清样本即可检测 9900 到 12,200 个特征，并在 50 分钟的梯度内鉴定来自 36 个亚类的 5842 种脂质。该方法可以很容易地适用于其他类型的生物样品，其中只有有限的体积可用。以及猪的脑脊液样本。该方法仅使用 1.0-2.5 μL 血清样本即可检测 9900 至 12,200 个特征，并在 50 分钟梯度内鉴定来自 36 个亚类的 5842 种脂质。该方法可以很容易地适用于其他类型的生物样品，其中只有有限的体积可用。