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Concept
Label-free quantification
Résumé
Label-free quantification is a method in mass spectrometry that aims to determine the relative amount of proteins in two or more biological samples. Unlike other methods for protein quantification, label-free quantification does not use a stable isotope containing compound to chemically bind to and thus label the protein.
Label-free quantification may be based on precursor signal intensity or on spectral counting.
The first method is useful when applied to high precision mass spectra, such as those obtained using the new generation of time-of-flight (ToF), fourier transform ion cyclotron resonance (FTICR), or Orbitrap mass analyzers. The high-resolution power facilitates the extraction of peptide signals on the MS1 level and thus uncouples the quantification from the identification process.
In contrast, spectral counting simply counts the number of spectra identified for a given peptide in different biological samples and then integrates the results for all measured peptides of the protein(s) that are quantified.
The computational framework of label free approach includes detecting peptides, matching the corresponding peptides across multiple LC-MS data, selecting discriminatory peptides.
Intact protein expression spectrometry (IPEx) is a label-free quantification approach in mass spectrometry under development by the analytical chemistry group at the United States Food and Drug Administration Center for Food Safety and Applied Nutrition and elsewhere. Intact proteins are analyzed by an LCMS instrument, usually a quadrupole time-of-flight in profile mode, and the full protein profile is determined and quantified using data reduction software. Early results are very encouraging. In one study, two groups of treatment replicates from mammalian samples (different organisms with similar treatment histories, but not technical replicates) show dozens of low CV protein biomarkers, suggesting that IPEx is a viable technology for studying protein expression.
Source officielle
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