GEN-MKT-18-7897-A
May 11, 2016 | Biopharma, Blogs | 0 comments
Fast LC-MS acquisition and automated data processing will help you speed up peptide mapping of your biotherapeutic, including critical disulfide bond and post-translational modification characterization. SCIEX helps you untangle the complexity of disulfide bonds, speeding up your characterization process.
See the full article by downloading the compendium >
Accurate disulfide bond mapping is essential for correctly establishing structure-function relationships as well as for monitoring the structural integrity of recombinant monoclonal antibodies (mAbs) throughout their production. Inappropriate disulfide bonds can affect a mAb’s stability, potency, aggregation, and may also signal errors in the cell culture or purification process. By following a biotherapeutic’s disulfide patterns over time, manufacturers can quickly detect production problems and then correct them as early as possible.
Correctly assigning disulfide bonds in a mAb, or ADC can be challenging and time-consuming due to the heterogeneity, large size, and multiple cysteine residues found in these biomolecules. Traditional approaches for disulfide mapping are based on fast liquid chromatography-mass spectrometry (LC-MS) analysis; however, these methods can be inefficient and usually involve digestion with multiple enzymes, tedious data processing, and intensive manual inspection of chromatograms for the identification of any possible disulfide linkages.
As the biotherapeutics industry develops and expands, there is an urgent need for software tools that can rapidly facilitate and accelerate the higher-order structural characterization of biopharmaceutical products. To meet these requirements, SCIEX has developed BioPharmaView™ Software, a data processing suite that can reduce the complexity of the massive data sets generated during biotherapeutic analysis. BioPharmaView Software uses rapid processing tools to accelerate critical characterization assays–such as peptide mapping and disulfide bond identification– by automating peak assignments, simplifying data processing, and streamlining the reporting process.
Peak Assignment Reduces Time for Peptide Mapping ExperimentsTo identify and match peptides, BioPharmaView Software automatically scores b- and y- ions from the high-resolution MS/MS spectra; and then the highest scoring experimental peaks are compared to a list of theoretical masses automatically generated by the software. The peak assignment process is further enhanced by predicting the theoretical fragment ion masses for non-reduced, disulfide-linked peptides before comparison with experimental data. Including other criteria in the ion selection process–such as MS/MS scoring, multiple charge states, and a retention time (RT) filter–can also help reduce the time needed for peptide mapping experiments. This enables manufacturers to meet regulatory requirements more quickly during the production and marketing of a new biotherapeutic product.
The ResultsIn this article, we successfully developed an efficient and automated workflow that comprehensively identified every disulfide linkage in the Fab region of an mAb. The use of the high-speed, TripleTOF® LC-MS System contributed to time-savings during disulfide analysis by permitting accurate mass MS and MS/MS information to be collected simultaneously, providing the high-resolution data necessary for differentiating closely related species and confirming structural assignments. And by using BioPharmaView Software to process the dataset, identifying the location of five disulfide linkages in the Fab region of an mAb was completed in a fast and automated fashion.
See the results in the full article by downloading the Biologics Analytical Characterization Compendium >
In monoclonal antibody (mAb) development, assessment of purity and integrity of the protein in question is critical. CE‑SDS is the gold standard assay and is routinely run from analytical development through QC and lot release. It’s trusted because it consistently delivers quantitative, size‑based insight into purity and fragmentation, and it fits naturally into regulated environments.
In drug discovery and development, Metabolite Identification (Met ID) plays a critical role in understanding biotransformation pathways, ensuring safety, and meeting regulatory requirements. Advanced mass spectrometry techniques have revolutionized this process, particularly through electron-based fragmentation methods such as Electron Activated Dissociation (EAD) and Electron Transfer Dissociation (ETD). While both techniques leverage electron interactions to generate informative fragment ions, they differ significantly in mechanism, performance, and suitability for Met ID workflows.
In analytical laboratories, performance is not optional. Whether supporting regulated pharmaceutical workflows, high-throughput CRO operations, clinical reporting, or food and environmental testing, your mass spectrometry and capillary electrophoresis systems are critical to productivity, compliance, and scientific confidence.
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