Expression and characterization of a chimeric bispecific antibody against fibrin and against urokinase-type plasminogen activator

Hiroko Tada, Tomofumi Kurokawa, Takeshi Seita, Takeshi Watanabe, Susumu Iwasa

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

We have produced a chimeric bispecific antibody that has dual specificity of human fibrin and urokinase-type plasminogen activator (u-PA). Complementary DNAs for variable regions of both anti-fibrin and anti-u-PA antibodies were cloned from two murine hybridomas secreting respective antibodies using polymerase chain reaction (PCR) techniques, and joined to cDNAs for human constant regions to form chimeric antibody genes. Both of two expression vectors for chimeric anti-fibrin and chimeric anti-u-PA antibodies were sequentially introduced into Chinese hamster ovary cells, and stable transfectants secreting the chimeric bispecific antibody were obtained. The highest producer transfectant (SULF/C2-30) secreted high level (about 40 μg ml-1) of total chimeric IgG and about 2% of the IgG had the bispecific activity of binding with both antigens. The chimeric bispecific antibody was purified by a combination of affinity chromatographies employing antigen-coupled columns and 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 as the original murine bispecific antibody.

Original languageEnglish
Pages (from-to)157-174
Number of pages18
JournalJournal of biotechnology
Volume33
Issue number2
DOIs
Publication statusPublished - Mar 31 1994
Externally publishedYes

Keywords

  • Chimeric bispecific antibody
  • Clot lysis
  • Fibrin specificity
  • Polymerase chain reaction
  • Single chain urokinase-type plasminogen activator

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

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