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蛋白质消化、超滤和尺寸排除色谱法优选人血浆和血清外来体的分离
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Biochemistry
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JoVE Journal Biochemistry
Protein Digestion, Ultrafiltration, and Size Exclusion Chromatography to Optimize the Isolation of Exosomes from Human Blood Plasma and Serum
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蛋白质消化、超滤和尺寸排除色谱法优选人血浆和血清外来体的分离

DOI:
10.3791/57467-v

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09:22 min

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April 13, 2018

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, , , , , ,
1Department of Microbiology, Immunology and Pathology,Colorado State University, 2Department of Biological Sciences, Eck Institute for Global Health,University of Notre Dame

Chapters

  • 00:04Title
  • 01:06Preparation of Crude Sample (Either Plasma or Serum) by Pelleting Larger Vesicles and Cellular Debris
  • 02:04Digestion of Non-exosomal Blood Proteins and Removal of Small Proteins/Peptides by Centrifuge Ultrafiltration
  • 04:21Purification of Vesicles by Size Exclusion Chromatography (SEC)
  • 05:45Concentration of Purified Exosomes/Total Protein Quantification and Quantification/Sizing of Exosomes by Nanoparticle Tracking Analysis (NTA)
  • 07:09Results: Increased Vesicle Yield and Exosome-specific Markers After Purification
  • 08:15Conclusion

Summary

Automatic Translation

Automatic Translation

在这里, 我们提出了一个协议, 以净化外来体从血浆和血清与减少共同纯化的非 exosomal 血液蛋白。优化的协议包括超滤、蛋白酶处理和尺寸排除色谱。增强纯化外来体的好处下游分析, 包括更准确的定量的囊泡和蛋白质组特征。

Tags

Protein DigestionUltrafiltrationSize Exclusion ChromatographyExosome IsolationBlood PlasmaSerumProteomicsBiomarkersExtracellular VesiclesProteinase K

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