Quarles R H, Johnson D, Brady R O, Sternberger N H
Neurochem Res. 1981 Oct;6(10):1115-27. doi: 10.1007/BF00964417.
Rabbits were immunized with the myelin-associated glycoprotein (MAG) that had been purified from isolated rat brain myelin by selective extraction with lithium diiodosalicylate (LIS) and phenol followed by preparative SDS gel electrophoresis. Antibodies to MAG were detected qualitatively by immunodiffusion and quantitatively by a double antibody assay utilizing [3H]fucose-labeled MAG as antigen. The antisera were capable of precipitating between 300 and 500 micrograms of MAG/ml of serum under the conditions of the assay. Preincubation of the anti-MAG serum with other glycoproteins or glycolipids did not inhibit the precipitation of labeled MAG. Similarly, preincubation of the antiserum with LIS-phenol extracts of non-neural tissues did not inhibit the immune precipitation of MAG. The specificity of the antiserum was also indicated by the selective double antibody precipitation of MAG from solubilized whole myelin that contained a heterogeneous mixture of [3H]fucose-labeled glycoproteins. The antibodies to MAG were not effectively absorbed by whole brain homogenate or purified myelin, indicating that the antigenic site(s) is not accessible in the intact membranes, but can be exposed by treatment with detergent or partial purification. Low levels of antibodies reacting with MAG were detected in three rabbits with experimental allergic encephalomyelitis induced by injection of purified myelin in complete Freund's adjuvant.
用通过二碘水杨酸锂（LIS）和苯酚选择性提取，然后进行制备性SDS凝胶电泳从分离的大鼠脑髓鞘中纯化得到的髓鞘相关糖蛋白（MAG）对兔子进行免疫。通过免疫扩散定性检测抗MAG抗体，并通过以[3H]岩藻糖标记的MAG作为抗原的双抗体测定法定量检测。在该测定条件下，抗血清能够沉淀每毫升血清300至500微克的MAG。抗MAG血清与其他糖蛋白或糖脂预孵育不会抑制标记的MAG的沉淀。同样，抗血清与非神经组织的LIS-苯酚提取物预孵育也不会抑制MAG的免疫沉淀。抗血清的特异性还通过从含有[3H]岩藻糖标记的糖蛋白异质混合物的溶解的全髓鞘中选择性双抗体沉淀MAG来表明。抗MAG抗体不能被全脑匀浆或纯化的髓鞘有效吸收，这表明抗原位点在完整膜中不可及，但可以通过用去污剂处理或部分纯化来暴露。在三只通过在完全弗氏佐剂中注射纯化髓鞘诱导实验性变应性脑脊髓炎的兔子中检测到与MAG反应的低水平抗体。
