小鼠抗天花粉蛋白IgE型单抗的酶解及其Fab的制备

研究了Ｐａｐａｉｎ及Ｔｒｙｐｓｉｎ裂解小鼠抗天花粉蛋白ＩｇＥ单抗的条件及Ｆａｂ的制备。Ｐａｐａｉｎ和Ｔｒｙｐｓｉｎ两者都可产生Ｆ（ａｂ′）_２，分子量在１５０～１６０ｋＤ左右；经Ｐａｐａｉｎ裂解的主要产物中还有Ｆａｂ，分子量７２ｋＤ，可通过凝胶过滤获得纯的Ｆａｂ。而Ｔｒｙｐｓｉｎ裂解物经ＤＴＴ还原、碘乙酰胺烷化虽然也可得到Ｆａｂ′（ｔ），但不易纯化；可见，要制备Ｆａｂ以采用Ｐａｐａｉｎ裂解为好，而制备Ｆ（ａｂ′）_２则可采用Ｔｒｙｐｓｉｎ裂解。这二个酶的裂解速度是Ｔｒｙｐｓｉｎ大于Ｐａｐａｉｎ。

Proteolysis of Mouse Anti-TCS IgE Monoclonal Antibody and Preparation of its Fab

The proteolysis of mouse IgE with papain and trypsin were investigated. F(ab')_2 could be produced by these two enzymes and its molecular weighgt was about 150-160 kD;Fab was also presented in the papain digestion as a main product with the molecular weight of 72 kD,which may be purified by gel filtration. Fab' could be obtained from the trypsin digestion after being reduced with DTT and alkylated with iodoacetamide,but it was difficult to get relatively pure Fab'(t). The molecular weights of heavy chain fragments in F(ab')_2 and Fab were 53. 5 kD and 37. 5 kD respectively. Both of them also contained intact light chains(27 kD). In conclusion, papain is the enzyme of choice for preparing Fab,while trypsin may be used for the preparation of F(ab')_2.Rate of digestion of papain was slower than that of trypsin.