CHARACTERIZATION OF SUB-VISIBLE AND

VISIBLE ANITBODY AGGREGATES

 

Joshua Wayment1, Suvajyoti Guha1,2, Michael Tarlov1, Dean Ripple1, Michael Zachariah1,2

 

1 National Institute of Standards and Technology, Gaithersburg, MD 20899

2Department of Mechanical Engineering, Chemistry and Biochemistry, University of Maryland, College Park, MD 20742

 

 

In this study we employ electrospray differential mobility analysis (ES-DMA) and size exclusion chromatography (SEC) to characterize nanometer sized aggregates, and micro-flow imaging (MFI) and atomic force microscopy (AFM) to characterize sub-visible and visible macroscopic particles. An ES-DMA apparatus volatilizes particles into the gas phase and then classifies them based on the balancing of the drag and electrical forces and has the capability of characterizing oligomers of up to ~ 150 nm. Results are presented for four monoclonal antibodies of the IgG class in ammonium acetate at pH 7.0: a polyclonal antibody (IgG-A), Rituxan (RmAb), and a monoclonal antibody that is glycosylated (IgG-B) and deglycosylated (IgG–C) in the Fc region.  These four antibodies were suspended in 20 mmol/L ammonium acetate buffer systematically stressed at 70 oC for up to 120 minutes, and aggregate formation tracked using the above-mentioned techniques.