Molecular Weight Determination by LC-MS
The intact molecular weight has always been recognized as an important characteristic of biologics. Intact molecular weight analysis is used to demonstrate the protein/peptide products and verify the biologics. Measuring molecule weight is a requirement of the regulatory guidelines.
Yaohai Bio-Pharma offers intact molecular weight analysis under native, reduced, N-deglycosylated conditions. We employ Liquid Chromatography-Mass Spectrometry (LC-MS) to determine the intact molecular weight.
Regulatory Requirements for Molecular Weight
It is recommended in the ICH Q6 that “molecular weight (or size) is determined using size exclusion chromatography, SDS-polyacrylamide gel electrophoresis (under reducing and non-reducing conditions), mass-spectrometry (MS), and other appropriate techniques”.
Analytical Methods
| Analysis |
Methods |
| Molecular Weight |
LC-MS / Liquid Chromatography/Electrospray Mass Spectrometry (LC/ES-MS) of native, reduced, N-deglycosylated proteins or subunit |
Analytical Procedures
Intact Molecular Weight:
- Performing LC-MS on the native sample as received.
- LC-MS Data analysis.
Reduced Molecular Weight:
- Treating the protein sample with a suitable reducing agent;
- Performing LC-MS on the reduced protein;
- LC-MS Data analysis.
N-deglycosylated Molecular Weight:
- Removing the N-glycans with a suitable enzyme;
- Performing LC-MS on the N-deglycosylated protein;
- LC-MS Data analysis.
Reduced and N-deglycosylated Molecular Weight:
- Treating the protein sample with a suitable reducing agent;
- Removing the N-glycans with a suitable enzyme;
- Performing LC -MS on Reduced and N-deglycosylated protein;
- LC-MS Data analysis.
Protein Subunit Molecular Weight:
- Subunit Cleavage
- Performing LC-MS on Protein Subunit
- LC-MS Data Analysis
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