Department of Immunology, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.
Methods Mol Biol. 2020;2132:75-83. doi: 10.1007/978-1-0716-0430-4_8.
Siglecs are known to be bound and regulated by membrane molecules that display specific sialic acid-containing ligands and are present on the same cell (cis-ligands). Because of the low-affinity binding of Siglecs to the glycan ligands, conventional methods such as immunoprecipitation are not suitable for identification of Siglec cis-ligands. Here we describe efficient and specific labeling of cis-ligands of CD22 (also known as Siglec-2) on B lymphocytes by proximity labeling using tyramide. This method may also be applicable to labeling of cis-ligands of other Siglecs.
Siglecs 已知与膜分子结合并受其调控，这些膜分子上带有特定的含有唾液酸的配体，且位于同一细胞上（顺式配体）。由于 Siglecs 与糖链配体的亲和力低，因此免疫沉淀等传统方法并不适合鉴定 Siglec 的顺式配体。在这里，我们使用辣根过氧化物酶标记的酪胺（tyramide）通过邻近标记法，描述了在 B 淋巴细胞上 CD22（也称为 Siglec-2）的顺式配体的有效和特异性标记。该方法也可能适用于其他 Siglec 的顺式配体的标记。
