Antibody-Drug Conjugate (ADC) Drug-to-Antibody Ratio (DAR) Analysis in Serum Using IA-LC-HRMS

Library Number:
APNT135035288
Part Number:
720006673EN
Author(s):
Armand Ngounou and Mark Wrona
Source:
Waters
Content Type:
Application Notes
Content Subtype:
Application Notes
Related Products:
 
 
 
 

A Robust and reliable IA-UPLC-HRMS method for the qualitative analysis and quantification of T-DM1 is described.

We set out to measure T-DM1 DAR and drug load distribution in biological matrix at different pharmacokinetic-relevant concentrations using immunoaffinity ultraperformance liquid chromatography high resolution mass spectrometry (IA-UPLC-HRMS). Purification from mouse serum is performed using streptavidincoated magnetic beads. Antigen-specific capture, based on recombinant biotinylated HER2, was successful in retaining trastuzumab emtansine on the beads. T-DM1 was eluted under acidic conditions in an aqueous environment and separated by reversed-phase LC on an ACQUITY UPLC H-Class PLUS Bio System. The Xevo G2-XS QTof Mass Spectrometer enabled qualitative analysis and quantification of ADC samples (Figure 2). Herein we show that UPLC-QTof can monitor the DAR of the therapeutic and provide consistent, reproducible measurements even at low levels in matrix.


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