AI Article Synopsis
- Aiming to enhance the detection of blood-based biomarkers for Alzheimer's disease (AD), researchers developed a method for profiling undepleted human serum, addressing the challenge of abundant proteins that skew results.
- Using advanced techniques, including tandem-mass-tag labeling and extensive liquid chromatography, the study analyzed serum samples from both AD patients and controls, achieving a deep examination of protein presence.
- The results indicated significant differences in protein expression, highlighting notable changes in proteins related to mitochondria and fatty acid metabolism, suggesting potential pathways for further research in AD biomarkers.
Article Abstract
Background: Blood-based protein measurement is a routine practice for detecting biomarkers in human disease. Comprehensive profiling of blood/plasma/serum proteome is a challenge due to an extremely large dynamic range, as exemplified by a small subset of highly abundant proteins. Antibody-based depletion of these abundant proteins alleviates the problem but introduces experimental variations. We aimed to establish a method for direct profiling of undepleted human serum and apply the method toward biomarker discovery for Alzheimer's disease (AD), as AD is the most common form of dementia without available blood-based biomarkers in clinic.
Methods: We present an ultra-deep analysis of undepleted human serum proteome by combining the latest 11-plex tandem-mass-tag (TMT) labeling, exhaustive two-dimensional liquid chromatography (LC/LC) fractionation (the 1st LC: 3 h for 180 fractions, and the 2nd LC: 3 h gradient per fraction), coupled with high resolution tandem mass spectrometry (MS/MS). AD ( = 6) and control ( = 5) sera were analyzed in this pilot study. In addition, we implemented a multiplexed targeted LC-MS3 method (TOMAHAQ) for the validation of selected target proteins.
Results: The TMT-LC/LC-MS/MS platform is capable of analyzing 4826 protein components (4368 genes), covering at least 6 orders of magnitude in dynamic range, representing one of the deepest serum proteome analysis. We defined intra- and inter- group variability in the AD and control groups. Statistical analysis revealed differentially expressed proteins in AD (26 decreased and 4 increased). Notably, these altered proteins are enriched in the known pathways of mitochondria, fatty acid beta oxidation, and AGE/RAGE. Finally, we set up a TOMAHAQ method to confirm the decrease of PCK2 and AK2 in our AD samples.
Conclusions: Our results show an ultra-deep serum discovery study by TMT-LC/LC-MS/MS, and a validation experiment by TOMAHAQ targeted LC-MS3. The MS-based discovery and validation methods are of general use for biomarker discovery from complex biofluids (e.g. serum proteome). This pilot study also identified deregulated proteins, in particular proteins associated with mitochondrial function in the AD serum samples. These proteins may serve as novel AD candidate biomarkers.
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Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472024 | PMC |
| http://dx.doi.org/10.1186/s12014-019-9237-1 | DOI Listing |
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