Anti-A and anti-B hemagglutinin depletion during Cohn purification process of 5% immunoglobulin.

Abstract

Polyvalent immunoglobulin G (IgG) products obtained by fractionation of human plasma are widely used to treat a broad range of conditions, including immunodeficiency syndromes and autoimmune, inflammatory, and infectious diseases. For high-quality products and to minimize adverse events related to the use of intravenous IgG (IVIG) it is very important to perform detailed analyses of their components. One of these components, that in rare cases can cause severe hemolytic conditions, is the amount of hemagglutinins, natural antibodies that bind A and/or B (anti-A or -B) antigens present in red blood cells (RBCs). To characterize different IgG batches and to monitor the efficacy of the production procedure in the hemagglutinin reduction, a direct agglutination test (DAT) and a new flow cytometry (FC)-based assay were used for measuring the activity and the content of hemagglutinins in IgG samples obtained at different stages of the purification process. A total of 113 batches of 5% IVIG, produced in 2013 by Kedrion Biopharma, were analyzed for the ability to agglutinate RBCs by DAT. All batches tested were within the limits set by the European Pharmacopoeia. Three batches of 5% IVIG were analyzed for their hemagglutinin levels. The finished products and the production intermediates were evaluated by the DAT and the FC assay. A significant decrease of anti-A and anti-B titer after the Fraction (F)III precipitation was observed in all batches tested and an evaluation of the results obtained by the two methods was performed. This study shows that the hemagglutinin titer, accurately measured in a high number of 5% IVIG batches, is within the allowed limits for the DAT method. The specific production process employed, in particular the FIII precipitation step, successfully removes IgM and significantly reduces IgG class hemagglutinins.