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OBJECTIVE To establish an efficient method for quantification of adalimumab in human plasma based on UPLC-Q Exactive-Orbitrap MS platform combined with immuno-affinity enrichment strategy. METHODS Candidate surrogate peptides were screened by full MS/ddMS2 and the selective surrogate peptide was quantitatively analyzed by parallel reaction monitoring. Immunoglobulins and therapeutic antibodies in human plasma were extracted by magnetic beads coupled with protein A. The proteins were denatured at high temperature and digested by trypsin. Stable-isotope labeled adalimumab was used as internal standard. RESULTS The peptide GLEWVSAITWNSGHIDYADSVEGR in variable region of adalimumab heavy chain was selected as signature peptide, showing specificity and selectivity. Adalimumab demonstrated good correlation within the range of 1-32 μg·mL-1. Precision, accuracy and total error all met the verification requirements. Thirty-one clinical samples had measurable adalimumab concentrations by the established method and ELISA method, yielding a good correlation. The mean of difference between the two methods was 0.5 μg·mL-1. CONCLUSION The universal immuno-affinity mass spectrometry method established in the study is suitable for quantification of therapeutic antibodies, and can accurately and precisely determine adalimumab concentration, which provides a strategy for clinical monitoring.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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