A rapid and scalable method for selecting recombinant mouse monoclonal antibodies

Area of Science:

• Developmental Biology
• Immunology
• Molecular Biology

Background:

• Monoclonal antibodies (mAbs) are essential tools for cell and developmental biologists, particularly for wholemount fixed tissue analysis.
• Traditional methods for isolating mAbs are time-consuming and unpredictable, hindering research progress.
• There is a need for efficient and scalable techniques to produce high-affinity and selective mAbs.

Purpose of the Study:

• To develop a rapid, scalable, and predictable method for selecting and expressing recombinant mouse monoclonal antibodies.
• To generate mAbs equivalent to those produced by traditional hybridoma technology but with improved efficiency.
• To facilitate the distribution and use of valuable antibodies through plasmid-based expression.

Main Methods:

• Immunization of mice with pooled antigens to increase throughput.
• Cloning of rearranged light and heavy chains from limited hybridoma cells into a single expression plasmid.
• Selection of antibodies recognizing formalin-resistant epitopes using zebrafish cell surface receptor ectodomains expressed in mammalian cells.

Main Results:

• The new method significantly increased throughput in antibody selection and expression.
• Selected antibodies demonstrated expected staining patterns on wholemount fixed zebrafish embryos, validating their efficacy.
• Recombinant mAbs produced were comparable in quality to those from parental hybridomas.

Conclusions:

• This streamlined method enables the rapid selection of multiple high-quality monoclonal antibodies from a single immunized mouse.
• The plasmid-based distribution system simplifies antibody accessibility and application.
• The technique offers a significant advancement for antibody generation in cell and developmental biology research.