N-Glycolylneuraminic acid (Neu5Gc) is a dietary immunogenic sugar that metabolically incorporates into diverse glycoconjugates in humans. Anti-Neu5Gc antibodies are detected in all human sera, play a role in several inflammation-mediated pathologies and are potential diagnostics of such diseases. Currently, it is difficult to determine their levels in individual human samples due to diverse Neu5Gc-epitopes recognition. Here we describe a simple method for detection of human anti-Neu5Gc antibodies. We exploit the difference between two mouse models that differ only by Neu5Gc-presence (wild-type) or Neu5Gc-absence (Cmah-/- knockout). We characterize mouse serum from both strains by HPLC, lectin and mass-spectrometry. We then use Cmah-/- knockout sera to inhibit all non-Neu5Gc-reactivity followed by binding to wild-type sera to detect overall anti-Neu5Gc response in a single assay. We applied this methodology to characterize anti-Neu5Gc IgG and IgA in sera of Kawasaki disease (KD) patients at various stages compared to controls. KD is an acute childhood febrile disease characterized by inflammation of coronary arteries that untreated may lead to coronary artery aneurysms, thrombosis and myocardial infarction. This estimated response is comparable to the average of detailed anti-Neu5Gc IgG profile analyzed by a sialoglycan microarray. Both assays revealed an elevated response in acute KD patients with normal coronaries compared to patients with aneurysm or dilated coronaries.
- Copyright © 2013 by The American Association of Immunologists, Inc.