Schneider C, Newman R A, Sutherland D R, Asser U, Greaves M F
J Biol Chem. 1982 Sep 25;257(18):10766-9.
A method is described by which an immunoaffinity matrix was constructed by binding antibody directly or indirectly to protein A-Sepharose 4B followed by cross-linking of the complex with dimethyl pimelimidate. This allows optimal spatial orientation of antibodies and, thus, maximum antigen binding efficiency. The affinity matrices were stable to high and low pH buffers without any significant antibody loss. The optimal conditions of antibody saturation, cross-linker concentration, and elution system were established and affinity columns made with the monoclonal antibodies J5, W6/32, and OKT9 for one-step isolation of the common acute lymphoblastic leukemia-associated antigen, HLA-AB antigens, and transferrin receptor, respectively, from cell lysates. The same methodology was also applied to immobilize transferrin with polyvalent anti-transferrin antibodies. This was then used to isolate the transferrin receptor from cell lysates.
本文描述了一种方法，通过将抗体直接或间接结合到蛋白A-琼脂糖4B上，随后用二甲基庚二酸亚胺交联复合物来构建免疫亲和基质。这使得抗体具有最佳的空间取向，从而实现最大的抗原结合效率。亲和基质对高pH和低pH缓冲液均稳定，且抗体无明显损失。确定了抗体饱和、交联剂浓度和洗脱系统的最佳条件，并用单克隆抗体J5、W6/32和OKT9制备了亲和柱，分别用于从细胞裂解物中一步分离常见急性淋巴细胞白血病相关抗原、HLA-AB抗原和转铁蛋白受体。同样的方法也用于用多价抗转铁蛋白抗体固定转铁蛋白。然后用其从细胞裂解物中分离转铁蛋白受体。
