Comparison of two functional kappa light-chain transcripts amplified from a hybridoma

Three heavy‐chain and three kappa (κ)‐chain transcripts were amplified from hybridoma cells secreting a monoclonal antibody (mAb) against transferrin receptor. Sequence analysis via IMGT/V‐QUEST yielded the functional/aberrant prediction. Two functional κ‐chain transcripts, Vκ2 and Vκ3, and one functional VH1 were revealed. Comprehensive bioinformatics analyses including sequence alignment, phylogenetic tree, somatic hypermutation prediction, and three‐dimensional‐molecular structure modeling were used to predict the origin of the two κ‐chain transcripts. The results of bioinformatics analysis suggest that Vκ3 is derived from the myeloma partner of the hybridoma; Vκ2 is derived from B‐cell. Functional transcripts VH1 and Vκ2 and Vκ3 were then used to construct two chimeric antibodies chi‐C2 (Vκ2–VH1) and chi‐C3 (Vκ3–VH1), respectively. Antigen‐binding experiments showed that only chi‐C2 remained the same affinity as its parental mAb. Possible explanations for the coexistence of two functional κ‐chain transcripts and the different affinity of the two chimeric antibodies are discussed.