Automated, Generic Reagent and Ultratargeted 2D-LC-MS/MS Enabling Quantification of Biotherapeutics and Soluble Targets down to pg/mL Range in Serum.
Anal Chem
; 92(13): 9412-9420, 2020 07 07.
Article
en En
| MEDLINE
| ID: mdl-32510921
ABSTRACT
Mass spectrometry has recently emerged as a powerful analytical tool for the assessment of pharmacokinetics and biomarkers in drug development. Compared with ligand binding assays, a major advantage of mass spectrometry-based assays is that they are less dependent on high quality binding reagents, while a key limitation is the relatively lower sensitivity. To address the sensitivity issue, we have developed a generic reagent, ultratargeted two-dimensional liquid chromatography-tandem mass spectrometry (2D-LC-MS/MS) method which combines commercially available protein A affinity capture, targeted analyte isolation by 2D-LC, and targeted detection by multiple reaction monitoring (MRM). A targeted-2D-with-dilution configuration was designed to automate 2D-LC-MS/MS. This method was systematically evaluated using an anti-CD22 monoclonal antibody spiked into monkey and human serum, where lower limits of quantification (LLOQ) of 0.78 and 1.56 ng/mL were achieved, respectively. This represents an over 100-fold improvement in assay sensitivity compared to the conventional LC-MS/MS method. The performance of the method was further confirmed by analyzing another monoclonal antibody, bevacizumab, as well as a soluble antigen, circulating PD-L1. The results indicate that our method enables quantification of antibody therapeutics and antigen biomarkers in both clinical and nonclinical samples in the pg/mL to low ng/mL range. Protein A affinity capture was employed as a universal sample preparation procedure applicable to both full-length antibody therapeutics and antibody-antigen complexes. This novel method is also fully automated and proven to be highly robust for routine bioanalysis in drug development.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Espectrometría de Masas en Tándem
/
Anticuerpos Monoclonales
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Anal Chem
Año:
2020
Tipo del documento:
Article
País de afiliación:
Estados Unidos