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Updated: Jul 17, 2026
Generation of Recombinant Human IgG Monoclonal Antibodies from Immortalized Sorted B Cells
Published on: June 5, 2015
High-affinity monoclonal antibodies to PED/PEA-15 generated using 5 microg of DNA
K E Kilpatrick1, D P Danger, E A Hull-Ryde
1Department of Protein Sciences, Glaxo Welcome, Inc., Research Triangle Park, NC 27709,USA. kek17850@glaxowellcome.com
Abstract:
Class-switched, affinity-matured murine monoclonal antibody (MAb) producing cell lines reactive with PED/PEA-15 were generated and isolated in less than 4 weeks following polynucleotide immunizations using only 5 microg of DNA in conjunction with the Powderject gene gun. Somatic fusions of peripheral lymph node cells were performed 13 days after initiating delivery of DNA encoding the target antigen. The data presented demonstrates the rapid production, identification, and characterization of class-switched, affinity-matured MAbs that bind PED/PEA-15. The reported strategy enabled the rapid development of MAbs that are useful in enzyme-linked immunoadsorbent assay (ELISA), Western blotting, and immunoprecipitations.
Insights
Researchers rapidly developed monoclonal antibodies (MAbs) targeting PED/PEA-15 using DNA immunization and a gene gun. This efficient method produced high-affinity antibodies for various assays in under four weeks.
Area of Science:
- Immunology
- Molecular Biology
- Biotechnology
Background:
- Developing specific monoclonal antibodies (MAbs) is crucial for biological research and diagnostics.
- Traditional MAb production can be time-consuming and resource-intensive.
Purpose of the Study:
- To establish a rapid method for generating class-switched, affinity-matured MAbs against PED/PEA-15.
- To demonstrate the efficacy of polynucleotide immunization combined with a gene gun for accelerated MAb development.
Main Methods:
- Utilized polynucleotide immunization with a low DNA dose (5 microg) delivered via a Powderject gene gun.
- Performed somatic fusions of peripheral lymph node cells 13 days post-immunization initiation.
- Isolated and characterized monoclonal antibody producing cell lines.
Main Results:
- Successfully generated and isolated class-switched, affinity-matured murine MAbs reactive with PED/PEA-15.
- Achieved MAb production and characterization in less than 4 weeks.
- Demonstrated MAb utility in enzyme-linked immunoadsorbent assay (ELISA), Western blotting, and immunoprecipitation.
Conclusions:
- The described strategy enables rapid development of high-affinity MAbs.
- Polynucleotide immunization with gene gun delivery is an efficient approach for MAb generation.
- The generated MAbs are suitable for diverse immunological and biochemical applications.
