UHPLC Analysis of NIST mAb on Zenix SEC-300: Effect of Mobile Phase Salt Concentration and pH

UHPLC Analysis of NIST mAb on Zenix SEC-300

Conditions

instrument Thermo Ultimate 3000 BioRS
column Zenix SEC-300, 30 cm x 7.8 mm I.D., 3.0 µm
mobile phase   As indicated
flow rate 1.0 mL/min
column temp. 25 °C
detector UV, 280 nm
injection 3.0 µL
sample NIST mAb, 10 mg/mL, 12.5 mM histidine, pH 6.0

Description

Analysis Note Detecting and/or quantifying the amount of aggregates in a biotherapeutic is a critical quality attribute (CQA) that must be monitored and reported to regulatory bodies by biopharmaceutical companies.  Typically, size exclusion chromatography (SEC) is used to characterize degree of aggregation.  This application showcases the effects that two variables of the mobile phase can play in ability to resolve aggregated species from the main monoclonal antibody (mAb) peak: salt concentration and pH.  Using a slightly acidic mobile phase, a dilute phosphate buffer, and an elevated sodium chloride concentration resulted in an increase in resolution of the main antibody peak from an aggregated species, as compared to other mobile phase conditions.
Legal Information Zenix is a trademark of Sepax Technologies
LiChrosolv is a trademark of Merck KGaA, Darmstadt, Germany
Application No. MUR110618B

Materials

     
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