1 research outputs found
<i>DIGESTIF</i>: A Universal Quality Standard for the Control of Bottom-Up Proteomics Experiments
In bottom-up mass spectrometry-based
proteomics analyses, variability
at any step of the process, particularly during sample proteolysis,
directly affects the sensitivity, accuracy, and precision of peptide
detection and quantification. Currently, no generic internal standards
are available to control the quality of sample processing steps. This
makes it difficult to assess the comparability of MS proteomic data
obtained under different experimental conditions. Here, we describe
the design, synthesis, and validation of a universal protein standard,
called <i>DIGESTIF</i>, that can be added to any biological
sample. The <i>DIGESTIF</i> standard consists of a soluble
recombinant protein scaffold to which a set of 11 artificial peptides
(iRT peptides) with good ionization properties has been incorporated.
In the protein scaffold, the amino acids flanking iRT peptide cleavage
sites were selected either to favor or hinder protease cleavage. After
sample processing, the retention time and relative intensity pattern
of the released iRT peptides can be used to assess the quality of
sample workup, the extent of digestion, and the performance of the
LCāMS system. Thus, <i>DIGESTIF</i> can be used to
standardize a broad spectrum of applications, ranging from simple
replicate measurements to large-scale biomarker screening in biomedical
applications