4.6 Article

MALDI-MS-based biomarker analysis of extracellular vesicles from human lung carcinoma cells

Journal

RSC ADVANCES
Volume 11, Issue 41, Pages 25375-25380

Publisher

ROYAL SOC CHEMISTRY
DOI: 10.1039/d1ra04305f

Keywords

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Funding

  1. National Natural Science Foundation of China [61805271, 62074155]
  2. Scientific Instrument Developing Project of the Chinese Academy of Sciences [YJKYYQ20200034]
  3. Key-Area Research and Development Program of Guangdong Province [2019B020226004]
  4. Shenzhen Science and Technology Innovation Commission [JCYJ20170818154035069, KCXFZ202002011008124, JCYJ20200109115405930]
  5. Shenzhen Engineering Laboratory of Single-molecule Detection and Instrument Development [XMHT20190204002]
  6. CAS Key Laboratory of Health Informatics [2011DP173015]

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This study investigated and optimized critical experimental conditions for EV protein profiling using a human lung carcinoma cell line, combining differential ultracentrifugation and MALDI-TOF MS. Results showed that medium components and ultracentrifugation procedures play important roles in mass spectrometry detection. The study demonstrated the potential of MALDI-TOF MS-based EV analysis with optimized protocols for rapid screening of protein biomarkers associated with early cancer diagnosis.
Extracellular vesicles (EVs) are actively secreted by mammalian cells. They are increasingly recognized as promising circulating biomarkers of disease progression. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) is currently one of the most powerful techniques for the rapid analysis of biological samples, especially for discovering biomarkers for disease diagnosis and prognosis. It is unclear what cell culture medium components and EV isolation methods are suitable for MALDI-TOF MS analysis. Using a human lung carcinoma cell line (A549), we investigated and optimized the critical experimental conditions for EVs' protein profiling by combining differential ultracentrifugation and MALDI-TOF MS. The results demonstrated that medium components and ultracentrifugation procedures to extract EVs played important roles in MS detection. Compared with EV-depleted serum and normal serum medium, conditioned medium with 2% fetal bovine serum in this study maintained cell proliferation and displayed significant protein profiling of EVs. RPS27A (ribosomal protein), which plays an essential role in mRNA translation and ribosome assembly for the differentiation of cancer cells, was detected from the EVs of lung cancer cells associated with cancer cell migration and invasion. We also found the known tumor diagnosis marker, which is S100A10_S100 calcium-binding protein A10. Therefore, MALDI-TOF MS-based EV analysis with optimized experimental protocols can contribute to future development of rapid screening techniques of protein biomarkers associated with early cancer diagnosis.

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