Expression vectors for human-mouse chimeric antibodies.

Area of Science:

• Biotechnology
• Molecular Biology
• Immunology

Background:

• Recombinant antibody production is typically time-consuming and labor-intensive.
• Mammalian expression systems are crucial for producing complex glycoproteins like antibodies.

Purpose of the Study:

• To construct mammalian expression vectors for cost-effective, large-scale production of full-length chimeric antibodies.
• To facilitate the rapid generation of chimeric antibodies by enabling easy replacement of variable regions.

Main Methods:

• Construction of mammalian expression vectors encoding human constant and murine variable immunoglobulin regions.
• Utilized unique restriction sites for variable region exchange via PCR.
• Validated antibody expression and specificity using Western blot analysis.
• Employed methotrexate (MTX) selection for generating high-expressing clones.

Main Results:

• Successfully constructed and validated expression vectors for chimeric antibodies.
• Western blot confirmed the correct size, structure, and specificity of expressed chimeric antibodies.
• Demonstrated efficient production of a human-mouse chimeric antibody (HCAb) targeting colorectal cancer.
• Achieved high secretion yields (30 pg/cell/day) at 10⁻⁶ M MTX, enabling >100 mg/L production.

Conclusions:

• The developed vectors (pYR-GCEVH and pYR-GCEVL) significantly streamline chimeric antibody production.
• This method offers a convenient and rapid approach for high-level recombinant antibody manufacturing.
• The system is suitable for producing therapeutic antibodies and research tools.