Aim: Denosumab is a recombinant fully human IgG2 that has a high affinity and specificity for human RANKL. Commercially available RANKL labeled with an Fc fragment cannot be used to establish an indirect ELISA. To characterize denosumab pharmacokinetic a robust and accuracy method should be developed urgently. Results: In this study, an immunoaffinity enrichment method coupled with LC–MS/MS was established. The LC–MS/MS method acquired a linear range from 0.1 to 30 μg/ml. The intra- and inter-run precision (CV%) was within 11.5 and 10.5%, respectively. More importantly, the LC–MS/MS pharmacokinetic data were consistent with ELISA. Conclusion: This approach accelerated the quantification, reduced the costs and provided an alternative in case of lacking the special antigen to denosumab or a RANKL-biotinylated reagent.
Keywords:
- denosumab
- ELISA
- immunoaffinity enrichment
- LC–MS/MS
- RANKL