US 12,169,203 B2
Native microfluidic CE-MS analysis of antibody charge heterogeneity
Hongxia Wang, Briarcliff Manor, NY (US); Haibo Qiu, Hartsdale, NY (US); and Ning Li, New Canaan, CT (US)
Assigned to Regeneron Pharmaceuticals, Inc., Tarrytown, NY (US)
Filed by Regeneron Pharmaceuticals, Inc., Tarrytown, NY (US)
Filed on Jan. 30, 2020, as Appl. No. 16/777,230.
Claims priority of provisional application 62/799,331, filed on Jan. 31, 2019.
Claims priority of provisional application 62/851,365, filed on May 22, 2019.
Prior Publication US 2020/0249241 A1, Aug. 6, 2020
Int. Cl. G01N 33/68 (2006.01); G01N 1/40 (2006.01)
CPC G01N 33/6848 (2013.01) [G01N 1/4044 (2013.01)] 20 Claims
OG exemplary drawing
 
1. A method for detecting and/or discriminating between post-translational modification variants of an antibody of interest in a sample, comprising:
(a) subjecting a sample comprising one or more antibodies of interest to thermal stress to form a stressed sample;
(b) contacting said stressed sample with a protease to digest the sample into antibody fragments, wherein said antibody fragments comprise F(ab′)2 fragments and Fc fragments;
(c) separating said antibody fragments by molecular weight and/or charge in one or more capillaries using capillary electrophoresis under native conditions, wherein said capillary electrophoresis is in an integrated microfluidic platform and said native conditions preserve said F(ab′)2 fragments and said Fc fragments;
(d) eluting separated antibody fragments from the one or more capillaries; and
(e) determining the mass of the eluted antibody fragments by mass spectrometry analysis, thereby detecting and/or discriminating between post-translational modification variants of the antibody of interest,
wherein said capillary electrophoresis is coupled online to said mass spectrometer.