Department of Cell Research and Immunology, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, 69978, Israel.
Methods Mol Biol. 2020;2110:59-72. doi: 10.1007/978-1-0716-0255-3_4.
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.
异种移植的主要障碍之一是组织表达非人类哺乳动物碳水化合物 N-羟乙酰神经氨酸 (Neu5Gc)。这种 9 碳骨架酸性糖是 N-乙酰神经氨酸 (Neu5Ac) 的羟化形式，两者都是哺乳动物中最常见的两种唾液酸类型。人类 CMP-Neu5Ac 羟化酶编码基因的缺失决定了含有 Neu5Gc 的异种移植物的免疫原性。在这里，我们描述了一种使用亲和纯化的鸡抗 Neu5Gc IgY 检测哺乳动物来源组织中 Neu5Gc 的免疫组织化学方法。通过与游离 Neu5Gc 或 Neu5Gc-糖蛋白的竞争性抑制进一步证明了特异性，但不能与 Neu5Ac 或 Neu5Ac-糖蛋白。该方法可用于评估异种移植物潜在的 Neu5Gc 免疫原性。
