N-Glycolylneuraminic acid (Neu5Gc) is a common cell-surface ligand in animals which is not biosynthesized in humans, but it can be acquired in human tissue from dietary sources such as red meat. It is important to understand the relevance of this potentially immunogenic glycan on human health, and selective detection methods are needed that can distinguish Neu5Gc from its biosynthetic precursor that is common in humans (i.e., N-acetylneuraminic acid (Neu5Ac)). Here, we demonstrate that Neu5Gc can be selectively oxidized by an engineered variant of galactose oxidase without any reaction toward Neu5Ac. Oxidation of Neu5Gc itself allowed for the full spectroscopic characterization of the aldehyde product. In addition, we show that Neu5Gc is also oxidized when it is part of a typical animal oligosaccharide motif and when it is attached to a protein-linked N-glycan. Oxidation of Neu5Gc introduces bioorthogonal functionality that can be exclusively labeled. We demonstrate that in combination with sialidase-mediated hydrolysis, this two-enzyme system can provide a useful tool for the selective detection of Neu5Gc in complex biological samples such as the biopharmaceutical alpha acid glycoprotein.