Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Automated signature peptide approach for proteomics.

L Riggs1, C Sioma, F E Regnier

  • 1Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA.

Journal of Chromatography. A
|August 28, 2001
PubMed
Summary

This study automates multidimensional chromatography for proteomics, simplifying sample preparation and analysis. The new method significantly reduces analysis time for identifying signature peptides.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Ex vivo effects of insulin on the structural integrity of equine digital lamellae.

Equine veterinary journal·2018
Same author

Glycoproteomic profiling of serum peptides in canine lymphoma and transitional cell carcinoma.

Veterinary and comparative oncology·2009
Same author

Hot topic: Prepubertal ovariectomy alters the development of myoepithelial cells in the bovine mammary gland.

Journal of dairy science·2008
Same author

Spinning-disk self-referencing interferometry of antigen-antibody recognition.

Optics letters·2004
Same author

Fractionation of isotopically labeled peptides in quantitative proteomics.

Analytical chemistry·2001
Same author

Electrophoretically mediated microanalysis of beta-galactosidase on microchips.

Electrophoresis·2001

Area of Science:

  • Proteomics
  • Analytical Chemistry
  • Biochemistry

Background:

  • Automating complex proteomic workflows is crucial for high-throughput analysis.
  • Multidimensional chromatography offers enhanced separation but often requires manual steps.
  • Signature peptide analysis is vital for protein identification and quantification.

Purpose of the Study:

  • To develop and validate an automated multidimensional chromatographic method for proteomics.
  • To streamline the signature peptide approach using immobilized enzyme and affinity chromatography.
  • To reduce sample preparation time and increase overall analytical throughput.

Main Methods:

  • Automated reduction and alkylation of peptides in the autosampler.
  • On-column trypsin digestion using an immobilized enzyme column.

Related Experiment Videos

  • Selective enrichment of phosphorylated peptides using Ga(III)-loaded immobilized metal affinity chromatography (IMAC).
  • Further separation via reversed-phase liquid chromatography (RPLC) coupled to electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS).

    • Main Results:

    • Successful automation of the entire multidimensional chromatographic process.
    • Demonstrated selection of phosphorylated peptides from tryptic digests.
    • Achieved a total analysis time of under 2 hours in tandem column mode.
    • Potential for increased throughput with parallel column configurations.

    Conclusions:

    • The automated approach significantly simplifies and accelerates proteomic analysis.
    • This method is effective for the enrichment and analysis of specific peptide subsets, such as phosphopeptides.
    • The system offers flexibility for high-throughput proteomic studies with minimal manual intervention.