1. LC-MS/MS strategies for therapeutic antibodies and investigation into the quantitative impact of antidrug-antibodies.
- Author
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Ewles M, Mannu R, Fox C, Stanta J, Evans G, Goodwin L, Duffy J, Bell L, Estdale S, and Firth D
- Subjects
- Adalimumab blood, Adalimumab immunology, Adalimumab metabolism, Antibodies, Anti-Idiotypic metabolism, Antibodies, Monoclonal immunology, Antibodies, Monoclonal metabolism, Bacterial Proteins metabolism, Bevacizumab blood, Bevacizumab immunology, Bevacizumab metabolism, Chromatography, Affinity, Chromatography, High Pressure Liquid, Humans, Limit of Detection, Peptides analysis, Peptides isolation & purification, Receptor, ErbB-2 chemistry, Trastuzumab blood, Trastuzumab immunology, Trastuzumab metabolism, Trypsin metabolism, Antibodies, Anti-Idiotypic chemistry, Antibodies, Monoclonal blood, Tandem Mass Spectrometry
- Abstract
Aim: We aimed to establish novel, high-throughput LC-MS/MS strategies for quantification of monoclonal antibodies in human serum and examine the potential impact of antidrug antibodies., Methodology: We present two strategies using a thermally stable immobilized trypsin. The first strategy uses whole serum digestion and the second introduces Protein G enrichment to improve the selectivity. The impact of anti-trastuzumab antibodies on the methods was tested., Conclusion: Whole serum digestion has been validated for trastuzumab (LLOQ 0.25 µg/ml). Protein G enrichment has been validated for trastuzumab (LLOQ 0.1 µg/ml), bevacizumab (LLOQ 0.1 µg/ml) and adalimumab (LLOQ 0.25 µg/ml). We have shown the potential for anti-drug antibodies to impact on the quantification and we have subsequently established a strategy to overcome this impact where total quantification is desired.
- Published
- 2016
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