Characterization of cerebrospinal fluid via data-independent acquisition mass spectrometry

ABSTRACT: Cerebrospinal fluid (CSF) is in direct contact with the brain and serves as a valuable specimen to examine diseases of the central nervous system through analyzing its components. These include the analysis of metabolites, cells as well as proteins. For identifying new suitable diagnostic protein biomarkers bottom-up data-dependent acquisition (DDA) mass spectrometry-based approaches are most popular. Drawbacks of this method are stochastic and irreproducible precursor ion selection. Recently, data-independent acquisition (DIA) emerged as an alternative method. It overcomes several limitations of DDA, since it combines the benefits of DDA and targeted methods like selected reaction monitoring (SRM). We established a DIA method for in-depth proteome analysis of CSF. For this, four spectral libraries were generated with samples from native CSF (n=5) CSF fractionation (15 in total) and substantia nigra fractionation (54 in total) applying to CSF DIA of three replicates. The DDA and DIA methods for CSF were conducted with the same nanoLC parameters using a 180 minute gradient. Compared to a conventional DDA method, our DIA approach both increased the number of identified protein groups with 1574 compared to DDA with 648 over 50 % with a comprehensive spectral library (generated with DDA measurements from five native CSF and 54 substantia nigra fractions) and decreased the coefficient of variation to 6 %, compared to 11 % with a DDA method. We also could show that a sample specific spectral library generated only from native CSF increased the identification reproducibility from three DIA replicates to 90 % (77 % with a DDA method). Moreover, by utilizing a substantia nigra specific spectral library for CSF DIA over 60 brain-originated proteins could be identified compared to only eleven with DDA. In conclusion, the here presented optimized DIA method substantially outperforms DDA and could develop into a powerful tool for biomarker discovery in CSF.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Brain

SUBMITTER: Katalin Barkovits  

LAB HEAD: Katrin Marcus

PROVIDER: PXD010690 | Pride | 2018-09-17

REPOSITORIES: Pride