Clezardin P, Bougro G, McGregor J L
J Chromatogr. 1986 Feb 28;354:425-33. doi: 10.1016/s0021-9673(01)87043-6.
A tandem chromatographic procedure was used to isolate rapidly mouse IgM monoclonal antibodies. Mouse ascites fluids containing IgM monoclonal antibodies were first chromatographed on an anion-exchange Mono Q column connected to a fast protein liquid chromatography system. The IgM-rich fractions from the Mono Q column were then injected on a gel filtration Superose 6 column equilibrated with a low-ionic strength buffer and eluted with a high-ionic strength buffer. Assessment of the purity of isolated IgM monoclonal antibodies was performed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis together with a Coomassie Brillant Blue R 250 staining technique. Assessment of the immunoreactivity of isolated IgM monoclonal antibodies was performed by a enzyme linked immunosorbent assay using a solid phase adsorbed antigen against which IgM monoclonal antibodies were directed. The chromatographic procedure described provides a new method for the rapid purification of mouse IgM monoclonal antibodies to a high degree of purity and in a immunoreactive state.
采用串联色谱法快速分离小鼠IgM单克隆抗体。首先将含有IgM单克隆抗体的小鼠腹水在连接到快速蛋白质液相色谱系统的阴离子交换Mono Q柱上进行色谱分离。然后将来自Mono Q柱的富含IgM的级分注入用低离子强度缓冲液平衡并用高离子强度缓冲液洗脱的凝胶过滤Superose 6柱上。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳结合考马斯亮蓝R 250染色技术对分离的IgM单克隆抗体的纯度进行评估。通过酶联免疫吸附测定法，使用针对IgM单克隆抗体的固相吸附抗原，对分离的IgM单克隆抗体的免疫反应性进行评估。所述色谱方法提供了一种新的方法，可将小鼠IgM单克隆抗体快速纯化至高度纯度并处于免疫反应状态。
