Practice question

Monoclonal antibodies originate immortalized hybridoma lines fusing single antibody-secreting B lymphocyte with myeloma partner preserving exactly one immunoglobulin gene rearrangement producing uniform paratope. Protocol begins immunization BALB/c mice purified antigen HER2 extracellular domain mixed Freund's adjuvant stimulating germinal center spleen. Four days after final booster splenocytes enriched plasma blasts expressing high-affinity surface IgG harvested. Fusion mediated PEG 1500 or electrofusion aligning cells pearl chains creates heterokaryons containing nuclei both parents tetraploid genome combining proliferative machinery myeloma P3X63Ag8U deficient HPRT and functional Ig heavy light chain variable regions B cell. Screening supernatants 10 days HAT medium ELISA identifies wells secreting antibody binding desired epitope single specificity affinity parent clone e.g., 10 nM. Cloning limiting dilution ensures monoclonality 0.5 cell per well. Resulting hybridoma inherits indefinite division activated c-myc autocrine IL-6 robust ER chaperone capacity folding continuous Ig secretion under Ig enhancers. Bioreactor expansion yields liters supernatant purified Protein A affinity chromatography producing uniform reagent defined sequence enabling therapeutics trastuzumab rituximab derived hybridoma technology now adapted recombinant CHO expression systems large-scale manufacturing.