Table 2.
Physicochemical and Biological Characterizations from the Comparability Studies of the Biosimilar Remsima74
| Characteristic | Attribute | Analytical Tool |
|---|---|---|
| Primary structure | Amino acid sequence | RP‐HPLC, LC‐ESI‐MS, LC‐ESI‐MS peptide mapping |
| Higher order structure | Disulfide structure | LC‐ESI‐MS peptide mapping |
| Free thiol analysis | Elman assay | |
| Secondary and tertiary structure | CD, FTIR, Antibody conformational array, X‐ray crystallography | |
| Thermal stability | DSC | |
| Purity | Monomer content | SEC‐HPLC, SEC‐MALS, SV‐AUC, CE‐SDS |
| Charge heterogeneity/amino acid modification | Charged isoforms | IEF, IEC‐HPLC |
| Deamidation/oxidation/C‐terminal variants | LC‐MS peptide mapping | |
| Glycosylation | N‐glycan analysis | LC‐MS |
| Glycosylation occurrence | CE‐SDS | |
| Oligosaccharide profile | HPLC | |
| Sialic acid analysis | HPAEC‐PAD | |
| Monosaccharide content (fucose, GlcNAc, galactose, and mannose) | HPAEC‐PAD | |
| Potency | Antigen and C1q binding | ELISA |
| FcRn binding | SPR | |
| Antigen neutralization | Cell‐based neutralization assay | |
| Apoptosis | Cell‐based apoptosis assay | |
| CDC | Cell‐based CDC assay |
CD, circular dichroism spectroscopy; CE‐SDS, capillary sodium dodecyl sulfate gel electrophoresis; DSC, differential scanning calorimetry; ELISA, enzyme‐linked immunosorbent assay; FTIR, Fourier transform infrared spectroscopy; GlcNAc, N‐acetylglucosamine; HPAEC‐PAD, anion exchange chromatography with the pulsed amperometric detection; IEF, isoelectric focusing; IEC‐HPLC, ion exchange chromatography; LC‐ESI‐MS, liquid chromatography electrospray ionization mass spectrometry; RP‐HPLC, reversed‐phase high‐performance liquid chromatography; SEC‐HPLC, size‐exclusion chromatography; SEC‐MALS, SEC‐multi angle light scattering; SPR, surface plasmon resonance; SV‐AUC, sedimentation velocity analytical ultracentrifugation.