UHPLC Analysis of Reduced Humira on BIOshell IgG 1000 Å C4

UHPLC Analysis of Reduced Humira on BIOshell IgG 1000 Å C4

Conditions

instrument Thermo Ultimate 3000 BioRS
column BIOshell IgG 1000 Å C4, 10 cm x 2.1 mm I.D., 2.7 µm
mobile phase [A] water (0.1% difluoroacetic acid); [B] acetonitrile (0.1% difluoroacetic acid)
gradient 0 to 100% B in 15 min
flow rate 0.3 mL/min
column temp. 75 °C
detector UV, 215 nm
injection 5.0 µL

Description

Analysis Note Humira (generic name adalimumab) is a monoclonal antibody (mAb) biotherapeutic used to treat arthritis, plaque psoriasis, Crohn's disease, and several other chronic illnesses.  The patent for Humira expried in 2016; therefore, there are several biosimilars of Humira that are being generated.  This application describes an HPLC method for characterizing Humira at the middle-up level.  After antibody reduction with dithiobutylamine, two sharp peaks are observed with the BIOshell IgG 1000 Å C4 column.  Exceptional resolution is achieved between the light and heavy chains with resolution of some minor variants around the peaks.
Legal Information BIOshell is a trademark of Sigma-Aldrich Co. LLC
LiChrosolv is a trademark of Merck KGaA, Darmstadt, Germany
SILu is a trademark of Sigma-Aldrich Co. LLC
Application No. MUR130418A

Materials

     
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