for protein identification & structural analysis
Characterize the protein in more details?
Need to know the protein isoforms in a protein batch?
Identify contaminating proteins from the host cell?
Alphalyse provides analytical characterization of your proteins.
Protein Structure Analyses
- Mass spectrometric peptide mapping with high sequence coverage
- N-terminal sequencing by Edman degradation
- C-terminal sequencing by MALDI ISD
- Mw determination of intact proteins by ESI MS & MALDI MS
- Quantitative Amino Acid Analysis
- Extinction coefficient analysis
- MRM quantification using LC MS/MS with isotope dilution standards
- 1D/2D gel electrophoresis with MS/MS protein identification
- iTRAQ multiplex labeling with nano LC MS/MS protein identification and relative quantification
- Host Cell Protein (HCP) detection & identification
The analyses are designed to support research, process development and analytical documentation. Analytical characterization of proteins is required by the ICH Q6B Guideline: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products, and the analysis results can be used in CMC documentation for regulatory authorities; EMA, FDA.
N-terminal Edman Sequencing
Determination of the amino acids sequence from the N-terminus using automated Edman degradation chemistry. Typically 5, 10 or 20 amino acids are sequenced for confirmation of expected N-terminal.
Molecular Weight Determination of Peptides and Proteins by MALDI MS and ESI MS
Intact protein Mw analysis for confirmation of expected Mw, and for detection of protein isoforms. Mw determination of unknown proteins and peptides.
N-terminal and C-terminal analysis by MALDI ISD top-down sequencing
Confirmation of full length protein sequence from both terminals. Detection of sequence truncations and terminal modifications.
1D SDS gels and 2D PAGE
Characterisation of protein Mw-isoforms and pI-isoforms. Separation and visualization of simple and complex protein mixtures for MS protein identification, N-terminal Edman sequencing and Western blotting.
Separation and purification of proteins and peptides based on hydrophobicity. Purity and impurity profile.
Peptide Mapping by MALDI MS and LC-ESI MS
Protease cleavage using trypsin, Glu-C, Asp-N, Lys-C, chymotrypsin followed by mass spectrometric peptide analysis. Confirmation of protein sequence. Detection and analysis of peptides with post-translational modifications.
Mass Spectrometric Protein Identification
Trypsin digestion, high sensitivity mass spectrometric peptide analysis 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 and trp oxidation, methylation, acetylation, free thiol, disulfide bonds.
Amino Acid Composition Analysis (AAA)
Hydrolysis and quantification of individual amino acids to determine amino acid composition and quantity of a protein sample.
Quantification and impurity assessment of proteins with known sequence.
Extinction Coefficient Determination
UV spectrophotometric analysis in combination with AA analysis for determination of the protein specific molar extinction coefficient.
Absolution Protein Quantification by LC-MS/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
Separation and identification of Host Cell Protein (HCPs) contaminants in protein drug substance by 1D SDS PAGE/ 2D PAGE, densitometric gel scan and mass spectrometric protein identification.