Which Biopharmaceuticals Can Be Analyzed by High-Resolution Mass Spectrometry for Molecular Weight?
- Intact antibody molecular weight (~150 kDa)
- Light-chain and heavy-chain molecular weights
- Molecular weight after deglycosylation
- Mass differences among glycosylation isomers
- Molecular weight changes before and after drug conjugation
- Average drug-to-antibody ratio (DAR)
- Content of unconjugated small-molecule drugs
- Accurate molecular weight of intact proteins
- Mass differences resulting from proteolytic cleavage or sequence variants
- Subtle mass shifts induced by post-translational modifications, such as phosphorylation and acetylation
- Molecular weights of linear and cyclic peptides
- Identification of amino acid point mutations
- Analysis of synthetic impurities and truncated species
- Molecular weight of intact oligonucleotides
- Minor mass differences resulting from chemical modifications, such as methylation or phosphorothioation
- Synthetic byproducts and truncated sequences
- Molecular weights of vaccine protein subunits
- Protein composition of recombinant virus-like particles (VLPs)
- Mass confirmation of high-abundance functional proteins within exosomes
High-resolution mass spectrometry (HRMS), owing to its exceptional mass accuracy, high sensitivity, and broad measurable mass range, has become an essential analytical tool in biopharmaceutical research and development. Beyond precise molecular weight determination of complex proteins, HRMS enables the resolution of subtle mass differences, providing critical support for structural characterization, impurity profiling, and quality control of biopharmaceutical products. Below, different categories of biopharmaceuticals are systematically examined to illustrate which molecular weights can be accurately measured using high-resolution mass spectrometry.
Monoclonal Antibodies
1. Measurable Items
2. Analytical Advantages
High-resolution mass spectrometry enables the interpretation of macromolecular spectral envelopes through deconvolution algorithms such as Xtract and MaxEnt, allowing accurate resolution of antibody variants with different glycosylation patterns. For IgG1 antibodies, for example, molecular weight differences arising from distinct sialylation or fucosylation modifications can be resolved at the sub-Dalton level.
Antibody-Drug Conjugates
1. Measurable Items
2. Application Scenarios
High-resolution mass spectrometry analysis of drug loading heterogeneity in ADCs supports the evaluation of DAR uniformity and overall product consistency.
Recombinant Proteins & Enzymes
1. Measurable Items
2. Analytical Advantages
High-resolution mass spectrometry plays an irreplaceable role in confirming the molecular integrity of recombinant proteins, particularly within the 10–100 kDa mass range, where it provides outstanding resolving power. High-precision qualitative characterization can also be achieved for recombinant proteins containing engineered domains or affinity tags.
Peptide Drugs
1. Measurable Items
2. Application Characteristics
Given their relatively low molecular weights (typically <10 kDa), peptide drugs are particularly well suited for High-resolution mass spectrometry analysis. In addition to accurate monoisotopic mass determination, HRMS enables clear discrimination of isomers, oligomers, and degradation products. Orbitrap-based platforms are especially advantageous for quality assessment of complex peptide formulations.
Nucleic Acid Drugs
1. Measurable Items
2. Technical Challenges And Solutions
Owing to their strong negative charge and susceptibility to degradation, oligonucleotides require optimized HRMS workflows incorporating specialized electrospray ionization conditions and advanced deconvolution software to achieve reliable molecular weight analysis.
Vaccine-Related Proteins And Exosome Proteomics
1. Measurable Items
2. Regulatory Trends In Biologics
As protein-based vaccines and exosome-derived products continue to emerge, regulatory agencies are imposing increasingly stringent requirements on protein composition and batch-to-batch consistency. High-confidence molecular weight determination by HRMS has become a core analytical component for compliant biopharmaceutical manufacturing.
From monoclonal antibodies and ADCs to peptides, oligonucleotides, exosomes, and vaccine proteins, high-resolution mass spectrometry spans the entire biopharmaceutical lifecycle, encompassing characterization, development, and quality control. Its central strengths include highly accurate molecular weight determination (error <5 ppm), robust isomer discrimination, and exceptional sensitivity to structural modifications and degradation. Leveraging advanced mass spectrometry platforms, including Orbitrap Exploris and Q Exactive series instruments, MtoZ Biolabs offers integrated, end-to-end solutions ranging from sample preparation to data deconvolution, enabling high-throughput and high-quality molecular weight analysis for biopharmaceutical products.
MtoZ Biolabs, an integrated chromatography and mass spectrometry (MS) services provider.
Related Services
How to order?
