A hybridoma-derived monoclonal antibody with high homology to the aberrant myeloma light chain

Autor: Boya Zhang, Anthony Berardi, Colin F. Greineder, Peter M. Tessier, Alec A. Desai, Henriette A. Remmer, Ghasidit Pornnoppadol
Rok vydání: 2021
Předmět:
Cell Lines
Immunoglobulin Variable Region
Myeloma
Artificial Gene Amplification and Extension
Polymerase Chain Reaction
law.invention
Hematologic Cancers and Related Disorders
Mice
Spectrum Analysis Techniques
law
Medicine and Health Sciences
Cloning
Molecular
Frameshift Mutation
Signal Amplification
Polymerase chain reaction
Multidisciplinary
Genes
Immunoglobulin
Antibodies
Monoclonal
Hematology
Flow Cytometry
Recombinant Proteins
Oncology
Spectrophotometry
Recombinant DNA
Medicine
Engineering and Technology
Biological Cultures
Cytophotometry
Antibody
Immunoglobulin Heavy Chains
Multiple Myeloma
Research Article
Cell Binding
Cell Physiology
DNA
Complementary
medicine.drug_class
Science
CHO Cells
Biology
Research and Analysis Methods
Immunoglobulin light chain
Monoclonal antibody
Cell Line
Frameshift mutation
Cricetulus
Antigen
Complementary DNA
medicine
Animals
Humans
Amino Acid Sequence
Myelomas and Lymphoproliferative Diseases
Molecular Biology Techniques
Molecular Biology
Immunohistochemistry Techniques
Cloning
Hybridomas
Tetraspanin 30
Cancers and Neoplasms
Biology and Life Sciences
Cell Biology
Molecular biology
Histochemistry and Cytochemistry Techniques
HEK293 Cells
Signal Processing
Immunologic Techniques
biology.protein
Immunoglobulin Light Chains
Zdroj: PLoS ONE
PLoS ONE, Vol 16, Iss 10, p e0252558 (2021)
DOI: 10.1101/2021.05.19.444780
Popis: The identification of antibody variable regions in the heavy (VH) and light (VL) chains from hybridomas is necessary for the production of recombinant, sequence-defined monoclonal antibodies (mAbs) and antibody derivatives. This process has received renewed attention in light of recent reports of hybridomas having unintended specificities due to the production of non-antigen specific heavy and/or light chains for the intended antigen. Here we report a surprising finding and potential pitfall in variable domain sequencing of an anti-human CD63 hybridoma. We amplified multiple VL genes from the hybridoma cDNA, including the well-known aberrant Sp2/0 myeloma VK and a unique, full-length VL. After finding that the unique VL failed to yield a functional antibody, we discovered an additional full-length sequence with surprising similarity (~95% sequence identify) to the non-translated myeloma kappa chain but with a correction of its key frameshift mutation. Expression of the recombinant mAb confirmed that this highly homologous sequence is the antigen-specific light chain. Our results highlight the complexity of PCR-based cloning of antibody genes and strategies useful for identification of correct sequences.
Databáze: OpenAIRE
Externí odkaz: