Protein Characterization

for structural analysis & protein identification

Can you document the structural identity of your protein, and identity of protein contaminants, in your purified protein batch?
Do you need characterisation of a therapeutic protein product?

Analytical characterization of proteins is required according to the ICH Q6B Guideline: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products.

Many questions and quality control issues are important when working with protein expression, purification and functional in-vitro and in-vivo assays. We provide analytical characterization of your protein product and protein contaminants to improve and speed up manufacturing process development. The results can be used as part of CMC documentation for regulatory authorities.

Applications:

• Mass spectrometric peptide mapping to get more than 90% sequence coverage of a specific protein
• Compare reference and batch protein preparations
• Confirm correct N-terminal processing from different cell clones
• Molecular weight determination to confirm full length expression and purification
• Host Cell Protein identification
• Drug substance variants analysis
• N- and C-terminal protein sequencing
  
  

Analytical Methods

N-terminal Edman Sequencing

Determination of the amino acids sequence from the N-terminus of the protein. Typically 6-15 amino acids for sequence confirmation.

Molecular Weight Determination of Peptides and Proteins by MALDI MS and ESI MS

Analysis of unknown proteins and peptides, or confirmation of expected sequence by Mw.

N- and C-terminal Sequencing by MALDI ISD

Confirmation of full length protein sequence and correct expression. Detection of sequence truncations and terminal modifications.

1D SDS gels and 2D PAGE

Characterisation of protein Mw-isoforms and pI-isoforms. Separation and purification of simple and complex protein mixtures for MS protein identification, N-terminal Edman sequencing and Western blotting.

Chromatography, RP-HPLC

Separation and purification of proteins based on hydrophobicity. Quantity and impurity profile.

Peptide Mapping by MALDI MS and LC-ESI MS

Protease cleavage by trypsin, Glu-C, Asp-N, Lys-C, chymotrypsin followed by mass spectrometry. Confirmation of protein sequence. Detection of post-translational modifications.

Mass Spectrometry Protein Identification Service

Mass spectrometric peptide mapping and database searching for identification of proteins in gel bands and chromatographic peaks.

Glycosylation/Phosporylation/Post-Translational Modification Analysis

Using affinity purifications, specific enzymes, chemical reactions combined with mass spectrometric peptide mapping and sequence correlation to detect, identify and compare modifications of different samples. Modifications include: deamidation, pyroglutatamate, met oxidation, methylation, acetylation, free thiol, disulfide bonds.

Amino Acid Composition Analysis (AAA)

Hydrolysis and quantification of individual amino acids to determine amino acid composition of protein/peptide.
Quantification and impurity assessment of proteins with known sequence.

Absolution Protein Quantification Assay by MS

Absolute quantification of specific protein in complex samples using stabile isotope labeled peptides (AQUA peptides) as internal standards for MRM measurements by LC-MS/MS.

Host Cell Protein (HCP) Identification Assay

Separation and identification of Host Cell Protein (HCPs) contaminants in protein drug substance by 1D SDS PAGE or 2D PAGE and mass spectrometric protein identification.