Incidence in plasma of low level antibodies against three xenotransplantation and immunotherapeutic glycan antigens

Antibodies against xeno-glycan antigens terminating with the saccharides Gal alpha, GalNAc alpha and Rha alpha are ubiquitous in human blood. Although originating as barriers to infection some of these naturally occurring complement-activating antibodies also contribute to disease processes, hinder xenotransplantation and have potential medical roles in immuno-oncotherapy. Because concentration of antibody is important in determining biological activity, there is a need to understand population variation in naturally occurring antibody levels, and to be able to rapidly and accurately determine levels in individuals. Xeno-glycan antigens in the form of function-spacer-lipid constructs were used to modify human red cells (kodecytes) to have on their surface micromolar equivalents of the xeno-glycan antigens Gal alpha 1-3Gal beta 1-4GlcNAc, GalNAc alpha 1-3Gal beta 1-4GlcNAc and Rha alpha. The methodology used was based on a previously validated kodecyte method used for quantifying IgM and IgG ABO human blood group antibodies in undiluted plasma. We tested plasma samples from 100 healthy individuals against these three different xeno-glycan kodecytes with each at three different loading concentrations of antigen to determine relative levels of these antibodies in human plasma. Sixty-one samples were also independently tested by enzyme immunoassay to correlate levels of anti-Gal alpha. Results demonstrate independence between antibody specificities and substantial variation between individuals in levels of these antibodies, with >92% of the population having medium or high levels of at least one specificity. However, of particular importance was that 5-8% of the population had low levels of both IgM and IgG to at least one specificity and these individuals would probably have a poor immediate response when challenged by the corresponding antigen.