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Sample Preparation and Relative Quantitation using Reductive Methylation of Amines for Peptidomics Studies
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Biochemistry
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JoVE Journal Biochemistry
Sample Preparation and Relative Quantitation using Reductive Methylation of Amines for Peptidomics Studies

Sample Preparation and Relative Quantitation using Reductive Methylation of Amines for Peptidomics Studies

DOI:
10.3791/62971-v

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08:00 min

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November 04, 2021

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, , ,
1Department of Biological and Environmental Sciences, Bioscience Institute,Sao Paulo State University (UNESP), 2Biodiversity of Coastal Environments Postgraduate Program, Bioscience Institute,Sao Paulo State University (UNESP)

Chapters

  • 00:04Introduction
  • 00:44Peptide Extraction: Cell Culture
  • 01:30Peptide Extraction: Animal Tissues
  • 03:39Peptide Quantification with Fluorescamine
  • 04:18Reductive Methylation of Amines Labeling
  • 05:58Results: Determining Labelled Peptides Using Mass Spectroscopy
  • 07:19Conclusion

Summary

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This article describes a sample preparation method based on heat-inactivation to preserve endogenous peptides avoiding degradation post-mortem, followed by relative quantitation using isotopic labeling plus LC-MS.

Tags

Sample PreparationRelative QuantitationReductive MethylationAminesPeptidomicsProtease InactivationIsotopic LabelingNeuroblastoma CellsZebrafish BrainCellular LysateUltrafiltrationSolid-phase Extraction

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