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Related Concept Videos

Proteomics01:33

Proteomics

7.3K
A proteome is the entire set of proteins that a cell type produces. We can study proteomes using the knowledge of genomes because genes code for mRNAs, and the mRNAs encode proteins. Although mRNA analysis is a step in the right direction, not all mRNAs are translated into proteins.
Proteomics is the study of proteomes' function. It involves the large-scale systematic study of the proteome to denote the protein complement expressed by a genome. Scientist Mark Wilkins coined the term...
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Ribosome Profiling

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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
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The technique...
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Related Experiment Video

Updated: Jul 5, 2025

Detection of Protein Ubiquitination Sites by Peptide Enrichment and Mass Spectrometry
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Detection of Protein Ubiquitination Sites by Peptide Enrichment and Mass Spectrometry

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Secretome processing for proteomics: A methods comparison.

Catarina Almeida-Marques1,2, Frank Rolfs1,2, Sander R Piersma1,2

  • 1Department Laboratory Medical Oncology, Amsterdam University Medical Center, Amsterdam, Netherlands.

Proteomics
|January 15, 2024
PubMed
Summary
This summary is machine-generated.

Acetone precipitation followed by in-solution digestion (ISD) is the best method for analyzing cancer secretome proteins. This approach enhances protein identification and reproducibility for biomarker discovery.

Keywords:
biomarkerin‐solution digestionproteomicssecretomesingle‐shot

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Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification

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Area of Science:

  • Proteomics
  • Cancer Biology
  • Biomarker Discovery

Background:

  • The cancer cell secretome offers valuable biomarkers due to its reflection of tumor-environment interactions.
  • High detectability of secretome proteins in biofluids makes them promising for diagnostics.

Purpose of the Study:

  • To evaluate six different workflows for processing cancer secretome samples.
  • To identify the optimal method for high-throughput secretome proteomics using single-shot mass spectrometry.

Main Methods:

  • Six secretome sample processing workflows were tested, involving protein concentration (ultrafiltration, acetone precipitation, lyophilization) and digestion (in-gel digestion, in-solution digestion).
  • Workflows were assessed based on protein identification numbers, unique identifications, reproducibility, and detection of small functional proteins (cytokines, chemokines, growth factors).
  • Single-shot nano liquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) was employed for analysis.

Main Results:

  • Acetone protein precipitation coupled with in-solution digestion (ISD) identified the highest number of proteins (2246).
  • This method also demonstrated superior reproducibility, with a correlation coefficient (r) of 0.94 and coefficient of variation (CV) of 19%.
  • In-solution digestion workflows, particularly with precipitation, showed improved identification of small, functionally important proteins.

Conclusions:

  • Acetone protein precipitation combined with in-solution digestion is the recommended method for high-throughput secretome proteomics.
  • This optimized workflow enhances the identification and quantification of cancer-associated proteins for biomarker discovery.