Size exclusion chromatography (SEC) is a common method for the separation of antibody monomer from dimer, aggregates, or degradation products on the basis of molecular size. YMC-Pack Diol-200 SEC columns with 2 μm exhibits reduced analysis time while achieving high resolution between monomer and Fab dimer. And with the new YMC-SEC MAB column an ideal analysis of fragments/degradation products of antibodies with high resolution can be achieved.
The characterisation of monoclonal antibodies (MAbs) is a major challenge in process monitoring and quality control. Size exclusion chromatography (SEC) is the standard method for analysis of fragments and also aggregates of MAbs in biopharmaceutical QC. The full characterization of Mab includes determination of Fab and Fc. For this application fragmentation to the MAb fragments is carried out using crystalline papain. When immunoglobulin are incubated with papain in the presence of a reducing agent, several peptide bonds above the hinge region are hydrolysed, producing three fragments of similar size (50 kDa)*:
- two monovalent Fab fragments
- one Fc fragment
The analysis of MAbs by SEC is typically performed under non-denaturating conditions at physiological pH range: 6 – 8. The commonly used buffer is phosphate buffer with NaCl.
>> Download the full Application Note as PDF
Newsletters
Receive the latest pathologist news, personalities, education, and career development – weekly to your inbox.
