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Peptide Mapping by LC-MS/MS Services
A setup fee may apply depending on number of samples and sample preparation time. The above quoted prices are based on analysis using our standardized methods, proven to work for the majority of antibodies and proteins. Method development options may be necessary for atypical proteins with unique properties.
Using peptide mapping, subtle changes to the primary structure of a protein or antibody can be identified that may affect the activity of the protein or the binding affinity. Peptide mapping by LC-MS/MS is one of the most powerful qualitative assays to confirm the primary sequence of proteins or antibodies. This service can be used to compare different lots of the same proteins, antibodies or biosimilars.
Service of peptide mapping analysis by LC-MS/MS includes:
A. The antibody/protein of interest is denatured, reduced, and digested with a proteolytic enzyme such as LysC or trypsin.
B. Optional: a second proteolytic enzyme is used to increase the sequence coverage of the protein.
C. The resulting peptide mixtures are separated on a reversed-phase HPLC. The chromatograph of UV absorbance is generated at UV 214 nm.
D. After HPLC separation, further analysis is performed by a high resolution mass spectrometry (Thermo Fisher Orbitrap Velos or Fusion).
E. A peptide map of the protein is generated for comparison with a reference protein.
F. The peaks are analyzed to the primary sequence.
G. Optional (two or more samples): a comparability evaluation is performed to assess lot-to-lot variation between proteins or to compare biosimilars.
- Confirmation of primary sequences (sequence coverage: 60-90% using one enzyme; > 98% using two or more enzymes).
- Glycoprofiling of N-linked oligosaccharides (heterogeneity of glycoforms & approximation of site occupancy).
- Identification & approx quantification of PTMs such as deamidation, oxidation, N-term pyroglutamate formation and C-term Lys processing (modified amino acid position and % modification can be detected).
- Similarity assessment between different lots.
- Comparability assessment for biosimilars.
Sample treatment: Denaturation, reduction/alkylation and enzymatic digestion followed by LC-MS or LC-MS/MS.