Specific Detection of Neu5Gc in Animal Tissues by Immunohistochemistry

Methods Mol Biol. 2020:2110:59-72. doi: 10.1007/978-1-0716-0255-3_4.

Abstract

One of the major obstacles in xenotransplantation is tissue expression of the non-human mammalian carbohydrate N-glycolylneuraminic acid (Neu5Gc). This 9-carbon backbone acidic sugar is the hydroxylated form of N-acetylneuraminic acid (Neu5Ac), and both constitute the two most common sialic acid types in mammals. Loss of CMP-Neu5Ac hydroxylase encoding gene in humans dictates the immunogenic nature of Neu5Gc-containing xenografts. Here we describe an immunohistochemistry method for the detection of Neu5Gc in mammalian-derived tissues using affinity-purified chicken anti-Neu5Gc IgY. Specificity is further demonstrated by competitive inhibition with free Neu5Gc or Neu5Gc-glycoproteins, but not with Neu5Ac or Neu5Ac-glycoproteins. This method can be used to evaluate potential Neu5Gc-immunogenicity of xenografts.

Keywords: Anti-carbohydrate antibodies; Immunohistochemistry; N-acetylneuraminic acid (Neu5Ac); N-glycolylneuraminic acid (Neu5Gc); Sialic acid; Xenotransplantation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomarkers
  • Humans
  • Immunohistochemistry / methods*
  • Liver / cytology
  • Liver / metabolism
  • Mammals
  • Mice
  • Neuraminic Acids / metabolism*
  • Organ Specificity
  • Sialic Acids / metabolism
  • Transplantation, Heterologous

Substances

  • Biomarkers
  • Neuraminic Acids
  • Sialic Acids
  • N-glycolylneuraminic acid