Beliaeva E L, Nikonenko B V, Sidorova E V
Biull Eksp Biol Med. 1979 Jul;88(7):116-9.
Two methods of isolating Fab- and Fc-fragments from mouse immunoglobulin G1 are presented. The first method involves fractionation of papain protein hydrolysate on a column with DEAE- (or DE-32)-cellulose adjusted with 0.005 M K-phosphate buffer, pH 8. The Fab-fragment was eluted from the column with the starting buffer. The Fc-fragment was eluted, with the buffer ionic strength being increased to 0.4 M. Another method involves protein fractionation on an ion exchanger adjusted with 0.004 M Tris-H3PO4 buffer, pH 8.5. All the protein was column bound. The Fab-fragment was eluted with 0.04 M Tris-buffer containing a 0.004 M mixture of K-phosphates, pH 8.6. The Fc-fragment was eluted, with ionic strength being raised to 0.4 M with phosphates. As none of the methods assures isolation of absolutely pure Fab- or Fc-fragments, it is requird that cross absorption of antisera with respective immunosorbents may be carried on in order to obtain monospecific antisera to these fragments.
本文介绍了两种从小鼠免疫球蛋白G1中分离Fab片段和Fc片段的方法。第一种方法是在装有经0.005M K-磷酸盐缓冲液（pH 8）调节的DEAE-（或DE-32）-纤维素的柱上对木瓜蛋白酶蛋白水解产物进行分级分离。Fab片段用起始缓冲液从柱上洗脱。Fc片段用离子强度增加到0.4M的缓冲液洗脱。另一种方法是在经0.004M Tris-H3PO4缓冲液（pH 8.5）调节的离子交换剂上进行蛋白质分级分离。所有蛋白质都结合在柱上。Fab片段用含有0.004M K-磷酸盐混合物的0.04M Tris缓冲液（pH 8.6）洗脱。Fc片段用磷酸盐将离子强度提高到0.4M后洗脱。由于没有一种方法能确保分离出绝对纯的Fab或Fc片段，因此需要用相应的免疫吸附剂对抗血清进行交叉吸收，以获得针对这些片段的单特异性抗血清。
