To characterize the L chain V region repertoire of IgG anti-Haemophilus influenzae type b capsular polysaccharide (Hib-PS) antibodies, clonal antibodies were purified from immune serum and internal amino acid sequences of V(K)II anti-Hib PS L chains obtained. We examined V(K)II L chains because it is the most common V(L) family expressed in the anti-Hib-PS response. Comparison of V(K)II amino acid sequences, including the entire CDR2 and CDR3 regions, of five anti-Hib-PS clonal antibodies from four unrelated individuals revealed complete identity with the exception of a single CDR3 residue from one antibody. When the sequence of these antibodies was compared with known V(K)II genes and myeloma proteins, it was found to be identical to the human V(K)II gene, A2, whose genomic sequence is presented here. In addition, all five of the V(K)II anti-Hib-PS antibodies examined contain an arginine inserted at the V-J junction. Finally, in contrast to the extraordinary homology of the V(K)II-encoded residues, there is variability in the JK gene utilization by these antibodies. These results demonstrate that the most common L chain V region in IgG anti-Hib-PS antibodies is the product of a single germ-line gene. The invariant arginine insertion suggests that this residue has an important role in Ag binding.