Quantitation of the monoclonal antibody Denosumab by bioassay and validated LC methods.
Int J Biol Macromol
; 119: 96-104, 2018 Nov.
Article
em En
| MEDLINE
| ID: mdl-30048724
The monoclonal antibody Denosumab (DmAb) is clinically used to treat osteoporosis and bone loss. We developed a bioassay based on the ability of DmAb to inhibit the effect of human receptor activator of nuclear factor-κB ligand (RANKL) to stimulate the formation of osteoclasts derived from RAW 264.7 cells. This bioassay was applied in conjunction with size exclusion high-performance liquid chromatography (SE-HPLC) and reversed-phase high-performance liquid chromatography (RP-HPLC) methods, with diode array detection (DAD), validated for the quantitation of this biotechnology-derived medicine. The SE-HPLC(DAD) method was carried out on a TSKGel G2000SWXL column and the mobile phase consisted of potassium phosphate buffer with sodium chloride, pHâ¯7.4. The gradient RP-HPLC(DAD) method was carried out on a Vydac 214TP C4 column at 60⯰C. The mobile phases consisted of 0.1% v/v trifluoroacetic acid (TFA) in water and 0.1% v/v TFA in acetonitrile. Calibration curves were linear over the concentration ranges 6-200⯵gâ¯mL-1 and 6-300⯵gâ¯mL-1 for the SE-HPLC(DAD) and RP-HPLC(DAD) methods respectively. The bioassay results correlated with the LC methods results, indicating the capabilities of these methods to quantitate DmAb, which will contribute to ensure the batch-to-batch consistency and efficacy of this biotherapeutic.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Bioensaio
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Cromatografia Líquida de Alta Pressão
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Denosumab
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Anticorpos Monoclonais
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
Idioma:
En
Revista:
Int J Biol Macromol
Ano de publicação:
2018
Tipo de documento:
Article
País de afiliação:
Brasil
País de publicação:
Holanda