Suppression of in vitro IgM rheumatoid factor production by diphtheria toxin interleukin 2 recombinant fusion protein (DAB 486IL-2) in patients with refractory rheumatoid arthritis.

OBJECTIVE: To investigate the effect of diphtheria toxin interleukin 2 recombinant fusion protein (DAB 486IL-2) on in vitro synthesis of immunoglobulin and rheumatoid factor (RF) in patients with severe refractory rheumatoid arthritis (RA) enrolled in a phase II, double blind, placebo controlled study. METHODS: Anticoagulated venous blood samples were obtained before (Day 1) and after (Day 28) intravenous infusion of either DAB 486IL-2 at 0.075 mg/kg/day (12 patients) or saline placebo (10 patients) on Days 1-5. Peripheral blood leukocytes (PBL) were prepared by density gradient centrifugation, cultured in the presence and absence of pokeweed mitogen (PWM) for one week, and culture supernatants assayed for immunoglobulins and IgM RF by ELISA. RESULTS: Compared to placebo treated patients, PWM induced IgM RF synthesis by PBL decreased after treatment with DAB 486IL-2 (p = 0.043). However, there was no apparent correlation with clinical improvement. PWM induced IgM, IgA, and IgG synthesis also tended to decrease, although the changes did not attain statistical significance. In contrast, PWM induced IgM RF, IgM, IgA, and IgG synthesis by PBL from patients treated with placebo tended to increase during the observation period. Spontaneous immunoglobulin and IgM RF production by PBL from either the DAB 486IL-2 or placebo patients remained stable. CONCLUSION: These observations raise the possibility that DAB 486IL-2 may diminish B cell function either directly or indirectly through effects on T cell function, but the change may not correspond to clinical response.

Polyethylene glycol precipitates of serum contain a large proportion of uncomplexed immunoglobulins and C3.

High pressure liquid chromatography (HPLC) was used to fractionate redissolved polyethylene glycol (PEG) precipitates isolated from the sera of normal volunteers and from patients with IgA nephropathy (IgAN) and systemic lupus erythematosus (SLE), 2 diseases characterized by elevated levels of circulating immune complexes. The individual fractions were analyzed by solid phase ELISA for IgA, IgM, C3, IgG, and complexes of IgG-IgA and IgG-C3. Although PEG precipitates were enriched for high molecular weight IgA and IgG (presumably bound within CIC), significant amounts of IgM, unbound IgG and C3 were also present. The quantities of the PEG-precipitable proteins did not correlate with their serum concentrations. IgG-IgA and IgG-C3 complexes were found in all precipitates examined, but the levels of complexes were higher in both patient groups. These results indicate that PEG precipitates a considerable quantity of proteins not bound in immune complexes. There appeared to be greater protein precipitation from sera of the patient groups compared to the amount precipitated from the normal sera. These results suggest that an understanding of the mechanism of PEG precipitation may be important in defining abnormalities in IgAN, SLE and perhaps other diseases characterized by elevated levels of CIC. In addition, the possibility of undetected CIC in PEG precipitable material must be considered.

Interleukin-2 diphtheria fusion protein (DAB486IL-2) in refractory rheumatoid arthritis. A double-blind, placebo-controlled trial with open-label extension.

OBJECTIVE: This pilot phase II, double-blind, placebo-controlled trial of 1 month duration, with a 2-3-month open-label extension, evaluated the safety, tolerability, biologic effects, and efficacy of interleukin-2 diphtheria fusion protein (DAB486IL-2) in refractory rheumatoid arthritis (RA). METHODS: Forty-five RA patients were enrolled in the trial, and were randomized, after a 3-4-week disease-modifying antirheumatic drug washout, to receive a daily intravenous dose of either DAB486IL-2 or placebo (saline) for 5 days. A blinded, third-party observer evaluated arthritis activity. Clinical response was defined as > or = 25% improvement in swollen and tender joints and > or = 25% improvement in at least 2 of 6 additional parameters. The double-blind phase was 4 weeks; placebo patients could cross over to receive open-label treatment for a maximum of 3 monthly DAB486IL-2 cycles. RESULTS: In the double-blind phase, 4 of 22 patients (18%) in the treated group and none in the placebo group (P = 0.05) met the criteria for clinical response. During the open-label treatment phase, 11 of 36 patients (31%) and 11 of 33 patients (33%) had a clinical response after completing 2 and 3 courses of DAB486IL-2, respectively. Adverse events included transient fever/chills (45%), nausea/vomiting (50%), elevated (< or = 3 x normal) transaminases (55%), and increased joint pain (45%). Twelve patients (8 placebo, 4 DAB486IL-2) did not complete 3 treatment cycles. No apparent differences were noted in CD4+ CD25+ cells of responders versus nonresponders, or of DAB486IL-2-treated versus placebo-treated patients. CONCLUSION: Clinical responses were noted in patients treated with DAB486IL-2 (18%) compared with placebo (0%) in the double-blind phase. In the open-label phase, 33% of patients completing 3 monthly DAB486IL-2 cycles had improvement in arthritis activity. Further studies of IL-2 diphtheria fusion proteins are warranted to elucidate factors that may predict clinical response and define mechanism(s) of action.

Activation of human neutrophils by surface-associated IgA is associated with the release of activated collagenase.

Neutrophils contain on their surface a receptor for the Fc portion of IgA. Cross-linking of this receptor in the fluid phase induces superoxide production and release of granule constituents, but the response to surface associated IgA has not been previously studied. Neutrophils incubated with surface-associated IgA (SAIgA) release significant amounts of activated collagenase in addition to the granule proteins myeloperoxidase and lactoferrin. This activation is associated with release of superoxide as well as hydrogen peroxide and hypochlorous acid. Although neutrophils incubated with soluble aggregates of IgA also release granule proteins and produce superoxide, soluble aggregates of IgA do not trigger the release of activated collagenase and do not generate hydrogen peroxide or hypochlorous acid. In summary, neutrophils activated by surface associated IgA respond differently than when cells are activated by soluble aggregates of IgA. These differences may be important in understanding the mechanisms of tissue injury in patients with inflammatory disorders.

Molecular characterization of monoclonal IgM derived from human B cell lines expressing the 4C9 rheumatoid factor associated idiotype.

Ten human monoclonal B cell lines that express the RF associated Id 4C9 were analyzed using an immunogenetic approach. Five of eight tested lines were also strongly positive for the 6B6.6 Id. We found that all the 4C9/6B6.6 positive lines expressed VkIIIa light chain genes. In contrast, 4C9 reactivity was also found on a cell line expressing a VkIIIb light chain gene and on a line expressing a V light chain gene. The two anti-Ids recognized a linear light chain determinant on Humkv328 encoded light chains but also a conformational determinant on Vg encoded light chains that appeared to be dependent on the presence of a heavy chain. Idiotypic reactivity occurred on both RF positive and RF negative antibodies. Within this idiotypic system, the basis for idiotypic reactivity and RF reactivity is complex, subject to both heavy and light chain gene usage and sensitive to small numbers of somatic mutations.

Shared idiotypes in mesangial deposits in IgA nephropathy are not disease-specific.

The antigenic specificity of the mesangial IgA in IgA nephropathy (IgAN) remains unknown. Because shared antigenic specificities may be reflected in the usage of shared idiotypes, we prepared five monoclonal anti-idiotypic antibodies (MoAbs) specific for the mesangial IgA eluted from the kidney of an IgAN patient. All five MoAbs reacted with the same idiotype, which proved to be of a public nature. Although the idiotype could be identified in the mesangial deposits of the majority of IgAN patients studied, it was not specific for the disease because it was also found in the glomerular deposits of other types of glomerulonephritis. The idiotype was also expressed in polyethylene glycol precipitates of sera and in pokeweed mitogen-induced plasma cells from both IgAN patients and healthy controls. The conclusion that no disease-specific idiotypes are present in the renal eluate was further supported by the failure to produce polyclonal anti-idiotypic antibodies by immunizing a rabbit with the eluted mesangial IgA. Our results support the concept that mesangial IgA deposits in IgAN are of a polyclonal nature.

Isolation, characterization and immunolocalization of a 53-kDal dentin sialoprotein (DSP).

We isolated a sialic-rich protein from rat dentin extracts and have named it dentin sialoprotein, DSP (formerly called 95K glycoprotein). DSP is rich in aspartic acid, glutamic acid, glycine and serine, but contains no cysteine or phosphate. The 30% carbohydrate content includes about 9% sialic acid and indicates that several N-glycosides and O-glycosides are present. Sedimentation equilibrium analysis gave a M(r) of 52,570. Based on this molecular weight we calculated that DSP contains about 350-amino acids and 75 monosaccharides. With automated Edman degradation the sequence of the first 8-amino acids was shown to be: Ile-Pro-Val-Pro-Gln-Leu-Val-Pro. The initial 3 residues of this sequence are identical to the first 3 in human osteopontin (OPN) and are closely similar to the Leu-Pro-Val sequences of OPN from other species, as well as at the beginning of bone acidic glycoprotein-75 (BAG-75). On Western immunoblots, purified polyclonal antibodies reacted only with DSP in dentin extracts and with none of the proteins from bone. Similarly, immunolocalization experiments showed the presence of DSP in dentin but not in enamel or alveolar bone. Along with immunohistochemical localization data reported elsewhere, these observations suggest that DSP may be an important marker for cells in the odontoblast lineage.

Abnormal galactosylation of serum IgG in patients with systemic lupus erythematosus and members of families with high frequency of autoimmune diseases.

Gas chromatographic carbohydrate analyses of IgG from 30 patients with idiopathic systemic lupus erythematosus (SLE) revealed lower content of galactose when compared to that in 36 controls of similar ages (mean +/- SD, 3.18 +/- 0.66 vs 3.82 +/- 0.41 galactose residues/mole of IgG, P < 0.001). Abnormal galactosylation was observed in 60% of SLE patients. Analyses of IgG from 58 members of five families, characterized by a high frequency of SLE and other autoimmune diseases and serological abnormalities, and 51 controls of similar age range revealed that IgG galactose deficiency was detectable not only in some members with clinical and serological abnormalities (P < or = 0.001), but also in those without evidence of autoimmune diseases or abnormal serologies (P < or = 0.001). These data indicate that abnormal galactosylation of IgG frequently occurs in asymptomatic members of families with a high frequency of SLE and other autoimmune diseases and suggests that this abnormality may be an indicator for the development of these diseases.

Restriction fragment length polymorphism analysis of HLA-DR, DQ, DP and C4 alleles in Caucasians with systemic lupus erythematosus.

HLA-DR, DQ, DP and C4 null alleles were determined by restriction fragment length polymorphism (RFLP) analysis in 60 Caucasian patients with systemic lupus erythematosus (SLE) and 66 controls. DR3 (DRw17) and DQw2.1 were increased in frequency in the patients with SLE associated with the presence of C4A null genes. HLA-DP alleles determined by RFLP analysis of genomic DNA as well as of PCR amplified DNA were not associated with SLE or any clinical or autoantibody subset thereof. No DR, DQ, or C4 null gene association was found with renal or neuropsychiatric involvement or nDNA antibodies (or levels thereof). These data suggest that the primary predisposition to SLE lies with HLA-DR or C4 null alleles, and not with HLA-DP.

Structural characterization of the second major cross-reactive idiotype group of human rheumatoid factors. Association with the VH4 gene family.

Rheumatoid factors (RF) are the most common type of functional antibodies among naturally occurring human monoclonal IgM proteins. A large subset of these autoantibodies use structurally homologous light chains of the kappa III subgroup, which bear the 6B6.6 cross-reactive idiotype (CRI). Although antibody binding activity requires both heavy and light chains, information about the heavy chains used by these autoantibodies is limited. To investigate these proteins, the murine monoclonal antibodies, 5-14 and 6-10, were generated by immunization with the heavy chains of the 6B6.6 CRI-positive RF, COR and LEW. These antiidiotypic antibodies reacted with 8 of 11 autoantibodies that coexpressed the 6B6.6 CRI. All 8 RF had heavy chains from the VH4 gene family, as assessed by reactivity with a VH4-specific primary sequence-dependent antibody. The same RF were also identified by the previously described murine monoclonal antiidiotype, LC1. Further experiments revealed that the LC1 antibody delineates a subfamily of VH4 heavy chains that is preferentially used in kappa III-6B6.6 CRI-positive IgM-RF. The cumulative data suggest that 13-22% of RF express both the kappa III-6B6.6 and VH4-LC1 CRI. These findings document that RF autoantibody activity requires specific VL-VH pairing, and that a subset of idiotypically related VH4 heavy chains is commonly expressed in disease-associated monoclonal IgM-RF.

Suppression of rheumatoid factor production by methotrexate in patients with rheumatoid arthritis. Evidence for differential influences of therapy and clinical status on IgM and IgA rheumatoid factor expression.

Suppression of rheumatoid factor (RF) production in rheumatoid arthritis (RA) has been variably attributed to the use of remittive agents per se or to clinical improvement associated with their use. There have been conflicting reports with regard to the influence of methotrexate (MTX) on serum RF levels in RA. We determined IgM-RF and IgA-RF levels in paired serum samples (obtained at study entry and completion) from RA patients enrolled in multicenter trials with the Cooperative Systematic Studies of Rheumatic Diseases program. After exclusion of the 14 IgM-RF-negative sera, there were samples from 30 MTX-treated patients and 52 placebo-treated patients. Changes in IgM-RF and IgA-RF levels were weakly associated with each other. Significant decreases in IgM-RF levels were observed in the MTX-treated patients, but not in the placebo group. These changes were most significant in the MTX-treated patients who improved clinically. There were significant decreases in IgA-RF levels at study completion among MTX-treated patients who had improved clinically and those who had not improved clinically, but not in the placebo group. The contributions of clinical improvement and MTX treatment to changes in serum IgM-RF and IgA-RF levels were examined using a logistic regression model. Changes in IgM-RF were strongly related to MTX treatment and, to a lesser extent, to clinical improvement; changes in IgA-RF were related only to MTX treatment. These results indicate that MTX treatment per se decreases both IgM-RF and IgA-RF levels, whereas clinical improvement correlates with decreased IgM-RF levels only.(ABSTRACT TRUNCATED AT 250 WORDS)

Dissociation of expression of two rheumatoid factor cross-reactive kappa L chain idiotopes in rheumatoid arthritis.

mAb 6B6.6 and 17.109 recognize two distinct kappa III L chain cross-reactive idiotopes (CRI) present on approximately 2/3 of IgM kappa rheumatoid factor (RF) paraproteins. To determine the distribution of these two CRI and their relationship to each other among polyclonal RF, sera from 86 RA patients and 49 controls were analyzed for the presence of 6B6.6- and 17.019-bearing RF by using sensitive solid phase ELISA. Levels of CRI(+) RF were estimated by using 6B6.6(+) and 17.019(+) RF standards. Detectable levels (greater than or equal to 195 ng/ml) of CRI(+) RF were rarely present in the control sera (8% for 6B6.6; 0% for 17.109), whereas 59% of RA sera contained measurable CRI(+) RF (48% for 6B6.6; 35% for 17.109; 21% for both). Where detected, CRI(+) RF were present in low concentrations (6B6.6: 1.21 +/- 1.56 micrograms/ml; 17.109: 1.20 +/- 1.15 micrograms/ml) and constituted a small fraction of the total IgM RF in these sera (6B6.6: 0.9 +/- 2.2%; 17.109: 0.8 +/- 0.9%). There was no correlation between either RF CRI and levels of IgM RF (r less than 0.1, p greater than 0.5). Levels of 6B6.6(+) RF did not correlate with 17.109(+) RF (r = -0.11, p = 0.47). In selected sera that contained both RF CRI, it was possible to selectively absorb 6B6.6(+) RF. Taken together, these data indicate the mutual independence of these two RF CRI among polyclonal RF and suggest the presence of distinct regulatory mechanisms governing their expression. Moreover, that these two CRI constitute a small proportion of polyclonal RF, in contrast to their striking predominance among monoclonal RF paraproteins, argues for the importance of other germline VL genes contributing to polyclonal RF production or the presence of extensive somatic mutation among polyclonal RF in RA.

Monoclonal antibody 6B6.6 defines a cross-reactive kappa light chain idiotope on human monoclonal and polyclonal rheumatoid factors.

Mouse monoclonal antibody (MAb) 6B6.6 was raised against a cross-reactive idiotope (CRI) present on the light chains of 2 human IgM paraproteins with rheumatoid factor (RF) activity. The MAb inhibited the IgG-binding activity of these proteins, and thus appears to react with an epitope located at or near the RF-binding site. Enzyme-linked immunosorbent assay (ELISA) and Western immunoblotting studies indicate that the 6B6.6 CRI is associated with kappa IIIa sub-subgroup light chains, is not related to the Wa, Po, and Bla RF cross-idiotypic specificities, and is clearly distinct from the kappa IIIb-associated CRI detected by MAb 17.109. Using an ELISA, we detected 6B6.6 CRI in 59% of 107 sera and 48% of 50 synovial fluids from patients with seropositive rheumatoid arthritis (RA). However, the quantities of CRI-positive RF were small, and the amount of CRI-positive RF did not correlate with the amount of IgM-RF. The 6B6.6 CRI was shown to occur primarily in the IgM fraction of RA sera by both chromatographic studies and isotype-specific ELISA, although small quantities appeared to be associated with IgA and IgG in some sera. The presence of 6B6.6 CRI on both monoclonal and polyclonal RF is consistent with the view that both are derived, at least in part, from a common gene pool. However, its occurrence in relatively low levels suggests that the number of germline genes encoding for RF is large or that extensive mutation occurs in the course of RF expression in RA.

Correlation of serologic indicators of inflammation with effectiveness of nonsteroidal antiinflammatory drug therapy in rheumatoid arthritis.

Forty-seven patients with rheumatoid arthritis (mean duration 5.7 years) who were receiving neither disease-modifying drugs nor corticosteroids were enrolled in a 12-week, multicenter study of the relationship between clinical and serologic measures of disease activity in patients taking nonsteroidal antiinflammatory drugs. After a 2-week drug washout period, patients received flurbiprofen (200 mg/day) or sustained-release ibuprofen (2,400 mg/day) for a 10-week trial. Clinical response was assessed biweekly using standard clinical parameters, including 50-foot walk time, tender joint score, duration of morning stiffness, and global assessment of disease activity and pain (by both the patient and the physician). Patients were classified as responders if there was greater than or equal to 30% improvement in at least 3 of the 4 clinical measures of disease activity. Thirty patients completed at least 8 weeks of therapy; there were 12 responders and 18 nonresponders. Of the laboratory parameters examined, the responders, but not the nonresponders, demonstrated significant reductions (from postwashout values) in levels of IgM rheumatoid factor and C-reactive protein (CRP), along with significant increases in the number of circulating lymphocytes and decreases in the number of circulating granulocytes (P less than or equal to 0.05). In contrast, the nonresponders demonstrated either no change or worsening of the laboratory correlates of disease activity. The responders also appeared to have more aggressive disease at baseline, with significantly more painful joints, greater 50-foot walk times, elevated CRP values, and elevated erythrocyte sedimentation rates (P less than or equal to 0.05). These data suggest that there is a subset of rheumatoid arthritis patients in whom clinical improvement with nonsteroidal antiinflammatory drug therapy is associated with significant reductions in IgM rheumatoid factor and CRP levels.

Occurrence of a rheumatoid factor cross-reactive kappa light-chain idiotope in rheumatoid arthritis families.

The occurrence of IgM rheumatoid factor (RF) and RF-associated kappa-III light-chain idiotope identified by monoclonal antibody 6B6.6 in the serum from 22 patients with rheumatoid arthritis (RA) and 68 relatives without connective tissue diseases in 15 families was determined by solid-phase enzyme-linked immunosorbent assays. Serum IgM RF was present in 19 RA patients from 12 families and 12 arthritis-free relatives of 4 families. It was not found in any of 9 spouses included in the study or in 44 of 45 unrelated healthy adult controls. RF-associated 6B6.6 idiotope was detected in 42% of the IgM RF(+) RA patients and in 50% of the IgM RF(+) arthritis-free relative, but not in the adult controls, spouses, and IgM RF(-) RA patients and relatives. It was present in one RA serum from each of 8 families and 6 sera from arthritis-free relatives of 2 families (5 of whom were from one family). Where present, the idiotope-positive RF represented only a small fraction of the serum IgM RF of the RA patients (0.1-2.1%) and relatives (1.5-14%). The increased frequency of IgM RF(+) individuals, with and without RA, in family groups suggests a genetic predisposition for expression of RF. The small proportion of RF bearing the 6B6.6 idiotope in both RA patients and unaffected family members supports the view that the number of germline genes encoding for RF is large or that extensive mutation occurs in the course of RF expression, whether idiopathic or associated with RA. In addition, nonuniform expression of the idiotope in RF within family groups indicates that the various clones of RF producing cells are to a large extent independently regulated.

DNA analysis of HLA-DR and DQ genes in American blacks with systemic lupus erythematosus.

We studied DNA polymorphisms of HLA-DR and DQ alleles in 63 American black patients with systemic lupus erythematosus (SLE). We found no HLA-DR beta, DQ alpha, or DQ beta restriction fragment length polymorphism (RFLP) or RFLP-determined DR or DQ specificity associated with SLE in either the patients or in 57 control subjects. DRw52b was positively associated with renal involvement and negatively associated with anti-nuclear RNP antibodies. Antibodies to Ro (SS-A) and La (SS-B) were associated with DR3(DRw17), DQw2.3. Early-onset SLE (less than or equal to 20 years of age) was associated with DRw8, and the frequency of neuropsychiatric involvement correlated negatively with a 3.7-kb Taq I DQ alpha RFLP. This suggests a role for DR and DQ genes in the clinical and serologic expression of SLE in American blacks.

Occurrence of polymeric IgA1 rheumatoid factor in the acquired immune deficiency syndrome.

The sera of 34 acquired immune deficiency syndrome (AIDS) patients and 20 healthy male homosexuals were examined for the presence of elevated levels of IgA and IgM rheumatoid factor (RF) and compared with results obtained with sera from 23 healthy laboratory volunteers. IgA RF levels were elevated (greater than 3 standard deviation units) in 9 of 34 (26%) patients with AIDS as compared to the panel of laboratory controls. Levels of IgM RF did not differ significantly in the AIDS patients and in the controls. There were no differences in levels of either IgA RF or IgM RF when the homosexual controls were compared with the laboratory volunteers. Sucrose-gradient ultracentrifugation experiments and assays using monoclonal reagents specific for IgA subclasses indicated that the IgA RF was predominantly of the polymeric configuration and restricted to the IgA1 subclass, respectively. Polyethylene glycol (PEG) precipitates of serum enriched for circulating immune complexes (CIC) were also assayed for the presence of IgA RF and IgM RF. Although levels of IgA RF in serum and in PEG precipitates did not correlate with levels of IgA- or IgA/IgG-containing CIC in AIDS patients, levels of IgA RF in both serum and CIC-enriched material were significantly elevated in the AIDS population when compared with the control panel. In contrast, levels of IgM RF in both serum and CIC-enriched material were low and not significantly different from those in healthy controls. These results indicate that both IgA-containing CIC and IgA RF occur in many AIDS patients and raise the possibility that IgA RF may contribute significantly to the formation of immune complexes in this disease.

Incidence of three cross-reactive idiotypes on human rheumatoid factor paraproteins.

The basis for rheumatoid factor (RF) production in autoimmune or lymphoproliferative diseases cannot be understood without defining the molecular factors that dictate RF structure and specificity. Recently three different mAb (6B6.6, 17.109, and G6) have been developed that define cross-reactive idiotypes (CRI) on intact L or H chains of human monoclonal RF cryoglobulins. However, the true incidence of these CRI among RF and their relationship to each other have not been delineated. In the present experiments, a panel of 163 randomly selected IgM paraproteins was evaluated for the expression of the two kappa L chain CRI, 6B6.6 and 17.109, and the H chain CRI, G6. Among the paraproteins with kappa L chains, 14% expressed the 17.109 CRI, and 9% expressed the 6B6.6 CRI. Both ELISA and Western immunoblotting experiments showed that the two L chain CRI were mutually exclusive. Anti-IgG activity was documented in 22 of the IgM-kappa paraproteins, among which mAb 6B6.6 reacted with 7 (32%) and mAb 17.109 with 6 (27%). Both CRI were expressed exclusively by L chains within the kappaIII variable gene subgroup. Although 17.109 CRI+ paraproteins had kappaIIIb L chains, none of the 6B6.6 CRI+ paraproteins possessed L chains with this kappa sub-subgroup specific Ag. The G6 CRI was found predominantly among RF paraproteins and was frequently yet not exclusively associated with the 17.109 CRI+ L chains. Additional experiments were performed on a panel of normal adult human sera and documented the presence of 6B6.6 and 17.109 CRI on a small percentage (0.1 to 2.0%) of IgM from most individuals. These data indicate that 1) the mAb 6B6.6 and 17.109 identify two major and distinct CRI among IgM-RF paraproteins, 2) both CRI are associated exclusively with kappaIII L chains, 3) kappaIIIb and kappaIII non-b L chains are equally prevalent among IgM-RF, 4) the G6 H chain CRI is frequently associated with 17.109 CRI+ L chains, but not with 6B6.6 CRI+ L chains, and 5) although the ability to make 6B6.6 and 17.109 CRI+ kappa L chains is common in humans, these CRI are present in low concentrations in normal IgM.