An immunoaffinity liquid chromatography–tandem mass spectrometry assay for the quantitation of matrix metalloproteinase 9 in mouse serum

An immunoaffinity liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed for the quantitation of the zinc endopeptidase matrix metalloproteinase 9 (MMP-9) from mouse serum. Sample preparation for the assay included magnetic bead-based enrichment using an MMP-9 antibody and wa...

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Bibliographic Details
Published in:Analytical biochemistry 2010-04, Vol.399 (2), p.202-210
Main Authors: Ocaña, Mireia Fernández, Neubert, Hendrik
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Chromatography, Liquid
Immunoaffinity
Immunomagnetic Separation - methods
Magnetic beads
Mass spectrometry
Matrix Metalloproteinase 9 - blood
Matrix Metalloproteinase 9 - isolation & purification
Mice
MMP-9
Peptides - analysis
Protein quantitation
Stable isotope-labeled peptide
Tandem Mass Spectrometry - methods
Trypsin - metabolism
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Summary:An immunoaffinity liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed for the quantitation of the zinc endopeptidase matrix metalloproteinase 9 (MMP-9) from mouse serum. Sample preparation for the assay included magnetic bead-based enrichment using an MMP-9 antibody and was performed in a 96-well plate format using a liquid-handling robotic platform. The surrogate peptide GSPLQGPFLTAR derived from MMP-9 by trypsin digestion was monitored using an on-line capillary flow trap–release chromatography setup incorporating a series of trap columns (C18, strong cation exchange, and another C18) prior to nanoflow chromatography and nanospray ionization with selected reaction monitoring (SRM) detection. The assay was fit-for-purpose validated and found to be accurate (
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2010.01.002