H. Tada et al., EXPRESSION AND CHARACTERIZATION OF A CHIMERIC BISPECIFIC ANTIBODY AGAINST FIBRIN AND AGAINST UROKINASE-TYPE PLASMINOGEN-ACTIVATOR, Journal of biotechnology, 33(2), 1994, pp. 157-174
We have produced a chimeric bispecific antibody that has dual specific
ity of human fibrin and urokinase-type plasminogen activator (u-PA). C
omplementary DNAs for variable regions of both anti-fibrin and anti-u-
PA antibodies were cloned from two murine hybridomas secreting respect
ive antibodies using polymerase chain reaction (PCR) techniques, and j
oined to cDNAs for human constant regions to form chimeric antibody ge
nes. Both of two expression vectors for chimeric anti-fibrin and chime
ric anti-u-PA antibodies were sequentially introduced into Chinese ham
ster ovary cells, and stable transfectants secreting the chimeric bisp
ecific antibody were obtained. The highest producer transfectant (SULF
/C2-30) secreted high level (about 40 mug ml-1) of total chimeric IgG
and about 2% of the IgG had the bispecific activity of binding with bo
th antigens. The chimeric bispecific antibody was purified by a combin
ation of affinity chromatographies employing antigen-coupled columns a
nd hydroxyapatite high-performance liquid chromatography. The purified
chimeric bispecific antibody significantly enhanced the thrombolytic
potency of single chain u-PA in an in vitro clot lysis assay as well a
s the original murine bispecific antibody.