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Peptidome analysis of cerebrospinal fluid by LC-MALDI MS.

Hölttä M, Zetterberg H, Mirgorodskaya E, Mattsson N, Blennow K, Gobom J - PLoS ONE (2012)

Bottom Line: Analysis of the extracts by offline LC-MALDI MS resulted in the detection of 3,000-4,000 peptide-like features.Out of these, 730 peptides were identified by MS/MS.The identified peptides were found to originate from 104 proteins, of which several have been reported to be involved in different disorders of the central nervous system.

View Article: PubMed Central - PubMed

Affiliation: Clinical Neurochemistry Laboratory, Institute of Neuroscience and Physiology, Department of Psychiatry and Neurochemistry, The Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden.

ABSTRACT
We report on the analysis of endogenous peptides in cerebrospinal fluid (CSF) by mass spectrometry. A method was developed for preparation of peptide extracts from CSF. Analysis of the extracts by offline LC-MALDI MS resulted in the detection of 3,000-4,000 peptide-like features. Out of these, 730 peptides were identified by MS/MS. The majority of these peptides have not been previously reported in CSF. The identified peptides were found to originate from 104 proteins, of which several have been reported to be involved in different disorders of the central nervous system. These results support the notion that CSF peptidomics may be viable complement to proteomics in the search of biomarkers of CNS disorders.

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Related in: MedlinePlus

LC-MS profiles of CSF peptides.The LC retention time is shown on the y-axis and the m/z on the x-axis. (a) Without sample pretreatment, 2,445 compounds were detected. (b) When adding 20% ACN prior to the ultrafiltration step, the number increased to 3,543.
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pone-0042555-g001: LC-MS profiles of CSF peptides.The LC retention time is shown on the y-axis and the m/z on the x-axis. (a) Without sample pretreatment, 2,445 compounds were detected. (b) When adding 20% ACN prior to the ultrafiltration step, the number increased to 3,543.

Mentions: From studies in plasma and CSF it is well known that many peptides bind to larger proteins and may therefore be retained in the ultrafiltration step [15], [26]–[28]. To improve the recovery of such peptides we investigated the effect of pretreating CSF samples with different concentrations of acetonitrile (ACN) and formic acid (FA), in order to dissociate the peptides from the carrier proteins prior to the ultrafiltration step. A CSF pool was divided into several 500 µl aliquots to which ACN or FA were added at different concentrations as described in the method section. The filtrates were analyzed by LC-MALDI MS and evaluated on the basis of number of detected compounds. Without any sample pretreatment, 2,445 compounds were detected (Figure 1 a). In a CSF sample treated with 20% ACN, the number of detected compounds increased to 3,543 (Figure 1 b). When the concentration of ACN was increased to 40% the amount of peptides in the flow through dropped significantly. A possible explanation for the observed decrease is that at this high ACN concentration, a large part of the sample proteins precipitate, resulting in co-precipitation of peptides. The highest numbers of compounds were detected when the CSF was incubated with 20% ACN. Addition of FA up to 5% to the samples also increased the number of detected compounds, however less than ACN. Increasing the FA concentration to 10% did not yield any additional improvement. Combining ACN and FA yielded poor results.


Peptidome analysis of cerebrospinal fluid by LC-MALDI MS.

Hölttä M, Zetterberg H, Mirgorodskaya E, Mattsson N, Blennow K, Gobom J - PLoS ONE (2012)

LC-MS profiles of CSF peptides.The LC retention time is shown on the y-axis and the m/z on the x-axis. (a) Without sample pretreatment, 2,445 compounds were detected. (b) When adding 20% ACN prior to the ultrafiltration step, the number increased to 3,543.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3412831&req=5

pone-0042555-g001: LC-MS profiles of CSF peptides.The LC retention time is shown on the y-axis and the m/z on the x-axis. (a) Without sample pretreatment, 2,445 compounds were detected. (b) When adding 20% ACN prior to the ultrafiltration step, the number increased to 3,543.
Mentions: From studies in plasma and CSF it is well known that many peptides bind to larger proteins and may therefore be retained in the ultrafiltration step [15], [26]–[28]. To improve the recovery of such peptides we investigated the effect of pretreating CSF samples with different concentrations of acetonitrile (ACN) and formic acid (FA), in order to dissociate the peptides from the carrier proteins prior to the ultrafiltration step. A CSF pool was divided into several 500 µl aliquots to which ACN or FA were added at different concentrations as described in the method section. The filtrates were analyzed by LC-MALDI MS and evaluated on the basis of number of detected compounds. Without any sample pretreatment, 2,445 compounds were detected (Figure 1 a). In a CSF sample treated with 20% ACN, the number of detected compounds increased to 3,543 (Figure 1 b). When the concentration of ACN was increased to 40% the amount of peptides in the flow through dropped significantly. A possible explanation for the observed decrease is that at this high ACN concentration, a large part of the sample proteins precipitate, resulting in co-precipitation of peptides. The highest numbers of compounds were detected when the CSF was incubated with 20% ACN. Addition of FA up to 5% to the samples also increased the number of detected compounds, however less than ACN. Increasing the FA concentration to 10% did not yield any additional improvement. Combining ACN and FA yielded poor results.

Bottom Line: Analysis of the extracts by offline LC-MALDI MS resulted in the detection of 3,000-4,000 peptide-like features.Out of these, 730 peptides were identified by MS/MS.The identified peptides were found to originate from 104 proteins, of which several have been reported to be involved in different disorders of the central nervous system.

View Article: PubMed Central - PubMed

Affiliation: Clinical Neurochemistry Laboratory, Institute of Neuroscience and Physiology, Department of Psychiatry and Neurochemistry, The Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden.

ABSTRACT
We report on the analysis of endogenous peptides in cerebrospinal fluid (CSF) by mass spectrometry. A method was developed for preparation of peptide extracts from CSF. Analysis of the extracts by offline LC-MALDI MS resulted in the detection of 3,000-4,000 peptide-like features. Out of these, 730 peptides were identified by MS/MS. The majority of these peptides have not been previously reported in CSF. The identified peptides were found to originate from 104 proteins, of which several have been reported to be involved in different disorders of the central nervous system. These results support the notion that CSF peptidomics may be viable complement to proteomics in the search of biomarkers of CNS disorders.

Show MeSH
Related in: MedlinePlus