We have compared the variable regions of 14 new IgM rheumatoid factors (RFs), produced in healthy human immunized donors (HIDs) with RFs originating from patients with rheumatoid arthritis (RA) and monoclonal Ig RFs (paraproteins or M-components, MC). Two groups with very restricted variable region structures were found. Twelve RFs (3 HID, 3 MC, and 6 RA) encoded by variable heavy (VH) chain germ-line genes with closest homology to DP-10 co-express the Kv325 variable light (VL) chain germ-line gene. These RFs have a remarkable restriction in the length (12-14 amino acids) and structure of the CDRH3. One HID RF has a CDRH3 only two amino acids different from the CDRH3 of a MC RF. Two sets of clonally related RFs, one from an RA patient and one from an HID, have CDRH3s that differ by only three amino acids. Five RFs (3 HID, 1 MC, and 1 RA) encoded by VH germ-line gene segments with closest homology to DP-54 all use the Kv328 VL germ-line gene combined to J kappa 1. Four are rearranged to the D21/9 D segment in the same reading frame, with CDRH3s of 16 to 17 amino acids. Three RFs (1 HID, 1 RA, and 1 MC) have CDRH3s differing by only three amino acids. The highly homologous V-regions in RFs from these two groups imply an initial selection to very similar, if not identical, epitopes. However, it remains to be seen whether somatic hypermutation alters the fine specificity of these autoantibodies.