Abstract
Human cerebrospinal fluid (CSF) is a sample of choice in the study of brain disorders. This biological fluid circulates in the brain and the spinal cord and contains tissue-specific proteins, indicative of health and disease conditions. Despite its potential as a valid source of biological markers, CSF remains largely understudied as compared to blood, in particular due to its more invasive way of sampling.
Challenges remain when performing proteomic analysis in clinical research studies. State-of-the-art mass spectrometry (MS) enables deep characterization of the human proteome. But some technical limitations are cardinal to be addressed, such as the capacity to routinely analyze large cohorts of samples. Importantly, a trade-off still needs to be made between the proteome coverage depth and the number of measured samples. In this context, we developed a scalable automated proteomic pipeline for the analysis of CSF. Because of its versatility, this workflow can be adapted to accommodate proteome coverage and/or sample throughput. It allows us to prepare and quantitatively analyze hundreds to thousands of CSF samples; it can also allow identification of more than 3000 proteins in a CSF sample when coupled with isoelectric focusing fractionation.
In this chapter, we describe an end-to-end pipeline for the proteomic analysis of CSF. The main steps of the sample preparation comprise spiking of a standard, protein digestion, isobaric labeling, and purification; these are performed in a 96-well plate format enabling automation. Depending on the targeted depth of the CSF proteome, optional analytical steps can be included, such as the removal of abundant proteins and sample pre-fractionation. Liquid chromatography tandem MS as well as data processing and analysis complete the pipeline.
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Macron, C., Núñez Galindo, A., Cominetti, O., Dayon, L. (2019). A Versatile Workflow for Cerebrospinal Fluid Proteomic Analysis with Mass Spectrometry: A Matter of Choice between Deep Coverage and Sample Throughput. In: Santamaría, E., Fernández-Irigoyen, J. (eds) Cerebrospinal Fluid (CSF) Proteomics. Methods in Molecular Biology, vol 2044. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9706-0_9
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DOI: https://doi.org/10.1007/978-1-4939-9706-0_9
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