Isotope Labeling

About 30,000 human genes are known to encode more than a million proteins, according to genomic research. Protein quantification, structural analysis, localization investigations, and post-translational modification detection are all parts of the field of proteomics, which is the study of proteins encoded by the genome. In proteomics, mass spectroscopy-based methods have become an effective tool. Regarding retention time, ionization effectiveness, and fragmentation mechanisms, stable isotope-labeled peptides (SIL peptides) are structurally and chemically identical to their endogenous counterparts. Therefore, both internal standards and template analytes are excellent choices. Incorporating amino acids having the required isotopes, such as deuterium (D), 13C, 15N, or 18O into the peptide during synthesis allows for the labeling of peptides with one or more hydrogen, carbon, nitrogen, or oxygen isotopes.

Utilizing SIL peptides as internal standards, the Absolute Quantification technique (AQUA) provides focused quantification of protein and post-translational changes in complicated protein mixtures. In a biological sample, the SIL peptide is added during or following protease digestion. Selective reaction monitoring (SRM) in a mass spectrometer is used to identify the heavy SIL peptide and the light endogenous peptide fragment that it is composed of. The quantity of endogenous protein can be determined using the known addition of SIL peptide and the intensity ratio of the two peptides.

Isotopic Labeling Chemistry

Isotopic Amino Acid Table

SIL Peptides in Structural Analysis

A potent method for examining the kinetics, structural details, and molecular interactions of biomolecules is nuclear magnetic resonance (NMR). The rate at which biomolecules relax as they separate from the target to which they are attached can be measured using this method. For NMR analyses of proteins, peptides labeled with D (spin of 1), 13C (spin 1/2), and 15N (spin 1/2) are appropriate. The mechanism of protein splicing, a post-translational autocatalytic process in which an intervening sequence, known as an intein, is excised from a host protein, the extein, is studied using a combination of segmental isotopic labeling and NMR spectroscopy.

High Quality SIL Peptides from CPC

High-quality peptides enriched with stable isotopes are necessary for structural studies and absolute quantification of a complex protein mixture at very low concentrations. Superior quality and high isotopic enrichment are promises made by the CPC Scientific SIL Peptide Custom Synthesis Service. In our cutting-edge peptide lab, these stable isotopic peptides are created using the most recent Fmoc solid-phase peptide technology. To determine the final purity and guarantee that our customers only receive the best peptides for absolute quantification and other investigations, all heavy isotope-labeled peptides go through rigorous analytical HPLC and mass spectrometric examination.

Peptides include two significant isotopes, 15N and 13C, in a certain order of amino acids. Each atom in the peptide can be found in several places and includes over 99% of an enriched isotope. For instance, a peptide containing a Leu amino acid that is isotopically tagged with one 15N and six 13C has a molecular weight that is seven units higher than the identical unmarked peptide.

SIL Peptide Applications

• Quantitative functional proteomics

• Quantification of proteins with post-translational modifications

• Study of protein structure

• Monitoring of protein expression

• Finding of biomarkers

• Pharmacokinetics

• Metabolomics

• Clinical biochemistry for monitoring of drugs and metabolites

• testing for doping

• Profiling of cell signals and verification of pathways

• Analysis of protein cross-links

SIL Peptide Modifications

• Phospho-Tyr, Ser, Thr (single or multiple)

• Sulfo-Tyr (single or multiple)

• Methylated Arg, Lys

• Chloro-Tyr

• Met-oxidized

• Pyroglutamic acid

Utilizing SIL peptides as internal standards, the Absolute Quantification technique (AQUA) provides focused quantification of protein and post-translational changes in complicated protein mixtures. In a biological sample, the SIL peptide is added during or following protease digestion. Selective reaction monitoring (SRM) in a mass spectrometer is used to identify the heavy SIL peptide and the light endogenous peptide fragment that it is composed of. The quantity of endogenous protein can be determined using the known addition of SIL peptide and the intensity ratio of the two peptides.