[Construction, expression and activity of a single chain antibody to gastric cancer cells].
OBJECTIVE
To construct single chain antibody from McAb 3H11, which has been successfully used in clinical radioimmunoimage, for its widel application in biological treatment of human cancer.


METHODS
Reversetranscription and polymerize chain reaction (RT-PCR) were used to clone the V region genes of McAb 3H11 (VH, VL). The cloned VH and VL were sequenced and recombined through a (Gly4Ser) 3 linker. The constructed single chain antibody gene was joined with glutathione-S-transferase gene (gst) and its fusion proteins (GS3H11) were expressed in E. coli. The activity of GS3H11 was characterized with competition experiments after its denature and renature treatment.


RESULTS
The recombinant GS3H11 was highly produced in inclusion bodies and the activity was recovered after being renatured. The competition experiments showed that GS3H11 could competitively inhibit the McAb 3H11 binding to gastric tumor cells MGC803. The inhibition was complete when the molecular ratio of GS3H11 to McAb 3H11 reached to 30:1. However, the GS3H11 did not interfere in binding of McAb PD4 and 3G9 to MGC803.


CONCLUSIONS
The specific binding activity of McAb 3H11 is retained in single chain antibody which is potentially useful for therapeutic application.
Shou
C
C
Department of Biochemistry and Molecular Biology, Beijing Institute for Cancer Research, Oncology School of Beijing Medical University.
Li
C
C
Meng
L
L
D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics; D000912:Antibodies, Neoplasm / Q000276:immunology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013274:Stomach Neoplasms / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins