BACKGROUND: Humans are genetically defective in synthesizing the common 
mammalian sialic acid N-glycolylneuraminic acid (Neu5Gc), but can metabolically 
incorporate it from dietary sources (particularly red meat and milk) into 
glycoproteins and glycolipids of human tumors, fetuses and some normal tissues. 
Metabolic incorporation of Neu5Gc from animal-derived cells and medium 
components also results in variable contamination of molecules and cells 
intended for human therapies. These Neu5Gc-incorporation phenomena are 
practically significant, because normal humans can have high levels of 
circulating anti-Neu5Gc antibodies. Thus, there is need for the sensitive and 
specific detection of Neu5Gc in human tissues and biotherapeutic products. 
Unlike monoclonal antibodies that recognize Neu5Gc only in the context of 
underlying structures, chicken immunoglobulin Y (IgY) polyclonal antibodies can 
recognize Neu5Gc in broader contexts. However, prior preparations of such 
antibodies (including our own) suffered from some non-specificity, as well as 
some cross-reactivity with the human sialic acid N-acetylneuraminic acid 
(Neu5Ac).
METHODOLOGY/PRINCIPAL FINDINGS: We have developed a novel affinity method 
utilizing sequential columns of immobilized human and chimpanzee serum 
sialoglycoproteins, followed by specific elution from the latter column by free 
Neu5Gc. The resulting mono-specific antibody shows no staining in tissues or 
cells from mice with a human-like defect in Neu5Gc production. It allows 
sensitive and specific detection of Neu5Gc in all underlying glycan structural 
contexts studied, and is applicable to immunohistochemical, enzyme-linked 
immunosorbent assay (ELISA), Western blot and flow cytometry analyses. 
Non-immune chicken IgY is used as a reliable negative control. We show that 
these approaches allow sensitive detection of Neu5Gc in human tissue samples and 
in some biotherapeutic products, and finally show an example of how Neu5Gc might 
be eliminated from such products, by using a human cell line grown under defined 
conditions.
CONCLUSIONS: We report a reliable antibody-based method for highly sensitive and 
specific detection of the non-human sialic acid Neu5Gc in human tissues and 
biotherapeutic products that has not been previously described.