Radioactive labeling of recombinant antibody fragments by phosphorylation using human casein kinase II and [gamma-P-32]-ATP

被引:38
作者
Neri, D
Petrul, H
Winter, G
Light, Y
Marais, R
Britton, KE
Creighton, AM
机构
[1] UNIV LONDON ST BARTHOLOMEWS HOSP & MED COLL,DEPT NUCL MED,LONDON EC1A 7BE,ENGLAND
[2] INST CANC RES,CTR CELL & MOL BIOL,LONDON SW3 6JB,ENGLAND
关键词
recombinant antibody phosphorylation; P-32; casein kinase II; gel-retardation;
D O I
10.1038/nbt0496-485
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
A wide range of antibody fragments can be expressed in bacteria and detected immunochemically via peptide tags. Using specially designed tags, we have developed a strategy for radiolabeling antibody fragments secreted from bacteria. Tagged antibody fragments were secreted either into the bacterial periplasm or the culture medium. The tag was not subject to proteolysis either in the broth or in human plasma. After affinity purification the antibody fragments were phosphorylated with [gamma-P-32]ATP and casein kinase II. The labeled fragments were used in a gel band-shift assay to measure antigen binding affinities. In contrast to non site-specific methods such as radioiodination, antibodies labeled with casein kinase II retain full immunoreactivity. Radioactively phosphorylated antibody fragments may have many other applications, including radioimmunoassays and radioimmunotherapy.
引用
收藏
页码:485 / 490
页数:6
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