Natural IgG Research Articles

Biological properties of a CD4 immunoadhesin

Molecular fusions of CD4, the receptor for human immunodeficiency virus (HIV), with immunoglobulin (termed CD4 immunoadhesins) possess both the gp120-binding and HIV-blocking properties of recombinant soluble CD4, and certain properties of IgG, notably long plasma half-life and Fc receptor binding. Here we show that a CD4 immunoadhesin can mediate antibody-dependent cell-mediated cytotoxicity (ADCC) towards HIV-infected cells, although, unlike natural anti-gp120 antibodies, it does not allow ADCC towards uninfected CD4-expressing cells that have bound soluble gp120 to the CD4 on their surface. In addition, CD4 immunoadhesin, like natural IgG molecules, is efficiently transferred across the placenta of a primate. These observations have implications for the therapeutic application of CD4 immunoadhesins, particularly in the area of perinatal transmission of HIV infection.

Anti-Ia treatment modulates specific and polyclonal antibody responses in Trypanosoma cruzi-infected mice

Experimental Chagas' disease—infection of mice with the protozoan parasite Trypanosoma cruzi—has been shown to increase the number of Ia-bearing cells in the spleen and the lymph nodes. The majority of these Ia-positive cells were Ig + and included in the large cell fraction of lymphoid organs from T. cruzi-infected animals indicating that they were activated B cells. These data are consistent with the polyclonal B-cell activation occurring during acute and chronic T. cruzi infection. The levels of secreted natural antibodies, of both IgM and IgG isotypes, were significantly increased in the sera of the infected animals. The present communication demonstrates that in vivo anti-Ia treatment of C3H HeJ mice infected with the CL strain of T. cruzi suppressed the polyclonal B-cell activation, affecting all the isotypes studied, including IgM, IgG2a and IgG2b, whose levels are predominantly increased during T. cruzi infection. In contrast to the decreased secretion of IgG autoantibodies, the levels of IgM autoantibodies were much less affected. The anti-Ia treatment totally abolished the specific anti-parasite response despite the fact that a pool of Ia-Ig positive cells remained after treatment.

IgG in murine intestinal secretions. Aging effect and possible physiological role.

To assess the role of, and age-associated alterations in, intraluminal IgG in the intestine, total IgG and subclass distribution in small intestinal perfusates and pilocarpine-stimulated intestinal secretions were measured under non-fasting conditions in three groups of BALB/c mice: 2-3, 12-14, and over 24 months old. Total intraluminal IgG and its subclasses (i.e. IgG1, IgG2a, and IgG2b) increased with age. However, the binding capacities of natural intraluminal IgG antibody (Ab) for outer membrane antigen (Ag) from normal habitant bacteria (Escherichia coli and Enterobacter agglomerans) were markedly reduced in the aged lavage samples as compared with such binding in samples from the other two age groups. The striking similarity to serum in the magnitude of the increase of intraluminal IgG and also in the distribution patterns of its subclasses in the small intestinal perfusates from all age groups suggests that the serum is a major source of the IgG. The intraluminal IgG may play some role in humoral immune protection under physiologic conditions of the gut lumen.

Natural occurring IgG and IgM antibodies to gram-negative bacteria and lipopolysaccharides in human sera

In sera of 28 non-infected elderly individuals, relationships between natural occurring antibody populations against O-antigen deficient lipopolysaccharides (J5LPS, ReLPS and Lipid A), a mixture of twelve O-antigen containing lipopolysaccharides (“mixed LPS”) and five different gram-negative bacterial strains, were assessed in a quantitative ELISA. Concomitant measurement of total IgG and IgM concentrations (in radial immunodiffusion assays—RIDA) allowed for the expression of IgG and IgM antibody concentrations per mg IgG.ml −1 and per mg IgM m −1 respectively. Statistically significant positive correlations were found between IgM antibody concentrations to all mentioned antigens for each possible combination ( r≥0·381; P<0·05). The strongest correlation was found between IgM concentrations to ReLPS and Lipid A ( r=0·852; P<0·001). IgG antibody concentrations to ReLPS correlated statistically significant with IgG antibody concentrations to Lipid A ( r=0·692; P<0·001), to J5LPS ( r=0·409; P<0·05) and to “mixed LPS” ( r=0·409; P<0·05). Beside the statistically significant correlations of IgG antibody concentrations to “mixed LPS” with ReLPS and with Lipid A ( r=0·528; P<0·001), IgG antibody concentrations to “mixed LPS” also correlated statistically significant with three gram-negative enterobacterial strains. These data indicate that the application of “mixed LPS” as a capture antigen for the selection of donor sera with a high IgG antibody concentration to this polyvalent antigen, eventually will yield a pooled serum which also contains relatively high IgG antibody concentrations to O-antigen deficient lipopolysaccharide preparations as well as enterobacterial strains.

Effects of in vitro and in vivo anticanine T lymphocyte and anticanine class II-specific monoclonal antibodies in the beagle.

Two murine anticanine lymphocyte monoclonal antibodies, designated T83 and B1F6, were assessed for (1) in vitro antiepitope activity to circulating lymphocyte subsets, their functional effects on lymphoproliferative assays and binding specificities to diverse cell suspensions and tissue sections; (2) in vivo effects after administration on cells expressing the target epitopes, lymphoproliferation, and allograft survival; and (3) the host immune response to the injected murine immunoglobulins. The monoclonal antibody T83 (IgG3) appeared to be specific for a T cell subset with an up-regulating function on lymphoproliferation. It caused a profound depletion of cells with the appropriate epitope after intravenous administration but it bound to lymphocyte membrane epitopes on some cells in peripheral blood that did not become depleted and putatively caused other modulating effects on lymphocyte function. The monoclonal antibody B1F6 (IgG2a, previously described) was immunologically specific in vitro for cells expressing class II major histocompatibility complex antigens. In the dog, this also consisted of 50% of T lymphocytes. After intravenous administration, there were functional effects similar to those of T83. A modest prolongation of survival of renal allografts was observed when both mAbs were used as the sole immunosuppressive agent. These studies also demonstrated the occurrence of natural canine antimurine IgG antibodies. Administration of either monoclonal antibody was followed by a rapid increase in the concentration of the antimouse antibody(s). We postulate that the presence of canine natural antimouse IgG markedly influenced the biologic effects of in vivo administered monoclonals.

Altered antibody isotype in cystic fibrosis: impaired natural antibody response to polysaccharide antigens.

Patients with cystic fibrosis (CF) have impaired natural (preinfection) IgG2 antibody responses to Pseudomonas aeruginosa lipopolysaccharide. To investigate the basis for this defect, we measured natural IgG and IgG1-4 antibody levels to Haemophilus influenzae type b polyribophosphate (PRP) and tetanus toxoid by enzyme-linked immunosorbent assay in 24 adult CF patients and 20 normal controls. Immunoglobulin heavy- and light-chain allotypes were determined on 146 Caucasian CF patients and 96 controls. The tetanus toxoid-specific IgG response was predominantly IgG1. CF and control subjects had similar IgG and IgG1 antibody levels. The PRP-specific IgG response was predominantly IgG2. In contrast to tetanus toxoid results, CF patients had lower geometric mean level of PRP-specific IgG compared to normal controls (p = 0.0036). ELISA results were confirmed by liquid-phase 3H-PRP-binding assay: CF patients had a geometric mean serum antibody level of 395 versus 922 ng/ml in controls (p = 0.0044). PRP-specific IgG2 levels were also depressed in CF patients (p = 0.03). CF patients had a lower prevalence of the A2m(2) allotype than the local racially matched control sample (p less than 0.025). Other allotype prevalences including G2m(n) and Km(1) were similar. Impaired IgG2 antibody responses to microbial polysaccharide surface antigens in CF patients might predispose them to persistent endobronchial infection and lead to production of nonopsonizing isotype responses. The potential role of A2m(2), coded for in the H chain locus on chromosome 14, is unknown, but could be related to mucosal IgA2 antibody responses.

IMPAIRED NATURAL IgG2 ANTIBODY (AB) RESPONSE TO POLYSACCHARIDE ANTIGENS AND DECREASED Km(1)–A2m(2) ALLOTYPES IN CYSTIC FIBROSIS (CF)

We recently described impaired IgG2 Ab responses to P. aeruginosa (PA) lipopolysaccharide in CF patients without PA infection (Pediatr Res 20:453, 1986). To investigate the basis for this defect, we measured natural IgG and IgG1-4 Ab levels to H. influenzae polyribophosphate (PRP) and tetanus toxoid (TT) by quantitative monoclonal Ab-based ELISA in 24 adult CF patients and 20 controls. Immunoglobulin allotypes were determined on 154 CF patients and 110 controls. The TT IgG Ab response was predominantly IgG1. CF and control subjects had similar TT IgG and IgG1 Ab levels. The PRP IgG Ab response was predominantly IgG2. In contrast to TT results, CF patients had subnormal levels of PRP IgG Ab compared to normal (geometric mean = 1.14 vs 2.32 meg/ml, respectively, 2 tailed p=0.004, Mann Whitney U test). PRP IgG2 levels were also depressed in CF patients (p=0.03). CF patients had a lower prevalence of Km(1) (p<0.025, X2 with continuity correction) and A2m(2) (p<0.05) than controls: G2m(n) and other allotype prevalences were similar. Impaired IgG2 Ab responses to certain polysaccharide-encapsulated microbes may predispose CF patients to endobronchial infection and lead to production of nonopsonizing isotype responses. A possible immunogenetic basis for impaired IgG2 Ab response is suggested by lower Km(l) allotype prevalence, while the role of A2m(2) is unknown.

The human natural anti-Gal IgG. III. The subtlety of immune tolerance in man as demonstrated by crossreactivity between natural anti-Gal and anti-B antibodies.

A well-defined antigen/antibody system was used to evaluate the effect of immune tolerance on the spectrum of specificities of natural antibodies. The antibody used in this study, anti-Gal, is a naturally occurring, polyclonal IgG that constitutes 1% of the circulating IgG in humans. We have previously shown that anti-Gal, purified from AB sera, specifically interacts with glycosphingolipids bearing a Gal alpha 1----3Gal epitope, but not with the closely related B antigen in which the penultimate galactose of the Gal alpha 1----3Gal epitope is fucosylated Gal alpha 1----3(Fuc alpha 1----2)Gal. This narrow specificity was assumed to be the result of an effective immune tolerance mechanism that prevents the expression of antibody clones that can recognize both the Gal alpha 1----3Gal and the self B epitopes. If the assumption that immune tolerance determines the range of anti-Gal specificity is correct, then anti-Gal from individuals lacking the B antigen (A and O blood types) would be expected to interact with both Gal alpha 1----3Gal and Gal alpha 1----3(Fuc alpha 1----2)Gal epitopes. In this study, anti-Gal from the serum of individuals of various blood types was purified by affinity chromatography on Gal alpha 1----3Gal adsorbent and tested for its reaction with the B antigen. Whereas anti-Gal from AB and B individuals only reacted with Gal alpha 1----3Gal epitopes, anti-Gal from A and O individuals reacted with both Gal alpha 1----3Gal and B epitopes. Furthermore, it was determined that the majority of anti-B reactivity in A and O individuals is in fact anti-Gal antibodies capable of recognizing both Gal alpha 1----3Gal and B epitopes. It can be concluded from these results that immune tolerance accurately controls the spectrum of natural antibody specificities by preventing the production of antibody clones that can interact with self antigens.

Differential recognition of oral indigenous bacteria by salivary immunoglobulins A and G.

Assuming that salivary immunity to indigenous microorganisms could develop, we assessed antibacterial reactivities of natural salivary antibodies in specific pathogen-free inbred mice. An ELISA was set up, using whole bacterial cells, to map reactivities of salivary IgA and IgG which accounted respectively for 91% and 8.7% of salivary Ig's in the BALB/c mouse. Representative strains of seven species from three genera (Lactobacilli, Staphylococci, and Streptococci), including major and minor components of the murine oral flora (38, 43, and 8%, respectively), were used to determine the presence and level of specific antibodies in individual saliva. It was verified that naturally occurring IgA antibodies can display diverse antibacterial reactivities. A characteristic profile emerged for salivary IgA where antibodies to Streptococcus faecalis predominate. Natural salivary IgG antibodies did not show the same reactivity pattern as IgA, anti-Lactobacilli and anti-Staphylococci reactivities being much less frequent in the salivary IgG repertoire. However, antibodies to S. faecalis occurred at the same high frequency for both isotypes (62-70% of the samples). Besides being species-specific, antibacterial reactivities were also found to be strain-specific. Broad variations in antibacterial titers were detected among individual mice under standardized experimental conditions. Present data thus suggest that the dynamics of salivary antibody production in the mouse reflect a differential natural sensitization of the secretory (IgA) versus the systemic (IgG) immune systems by distinct populations of indigenous bacteria.

Natural IgM and IgG Autoantibodies to Epidermal Keratins in Normal Human Sera. I: ELISA-Titration, Immunofluorescence Study

This paper presents a study of autoantibodies (autoAB) to keratins and to epidermis by a double approach associating a specific immunoenzymatic technique and immunofluorescence. The existence of natural autoAB to keratins in all normal human sera was asserted and the heterogeneity of natural autoAB to the epidermis explored. By a sensitive enzyme-linked immunosorbent assay we detected natural IgM and IgG autoAB to keratin polypeptides extracted from human plantar stratum corneum (SC) in 60 randomly selected normal human sera. The interindividual variation factors of their titers were about 100X in IgM and 50X in IgG antikeratin (AK) autoAB. The IgM and IgG AK autoAB titers varied independently. By a semiquantitative indirect immunofluorescence assay we detected in these sera IgM and IgG autoAB that labeled normal epidermis according to various morphologic patterns. The IgG autoAB labeled SC and suprabasal layers (SBL) in 57.4% of sera, SC in 20.4% and SBL in 7.4%. The IgM autoAB labeled SC and SBL in 52% of sera, SC in 24%, SC and SBL plus basal layer (BL) in 18%, and SBL in 2%. Like the titers, the patterns of IgM and IgG autoAB to epidermis were found to be unrelated. The IgG AK autoAB titers were found to significantly correlate only with the IgG autoAB directed to SC + SBL; the IgM AK autoAB titers only with the IgM autoAB directed to SC + SBL + BL. This showed that these patterns of labeling are typical for AK autoAB and that autoAB to SC, which could not be related to AK autoAB, exist in some normal sera. Antikeratin and antiepidermis IgM autoAB titers were found to be strongly correlated to total amounts of IgM assayed by radial immunodiffusion, indicating that the synthesis of these natural IgM autoAB vary in the same way as that of general IgM synthesis. For the AK and antiepidermis IgG autoAB, however, the same correlation to total serum IgG was found to be much weaker.

Human natural anti-alpha-galactosyl IgG. II. The specific recognition of alpha (1----3)-linked galactose residues.

A natural IgG antibody (anti-Gal) with alpha-galactosyl binding specificity has been found in large amounts (0.5 - 1.0% of serum IgG) in all individuals tested. It has been purified by affinity chromatography on a column of melibiose-Sepharose. In addition to its affinity for normal and pathological senescent human red cells, the antibody readily interacts with rabbit red blood cell (RRBC) glycolipids with alpha-galactosyl terminal residues. Two types (glycosidic linkages of 1----3 vs. 1----4) of rabbit red cells glycolipids with terminal alpha-galactosyl residues were tested for antibody binding. The antibody specifically bound to glycolipids with Gal alpha 1----3 terminal residues, and treatment of these glycolipids with alpha-galactosidase abolished binding. Hemagglutination inhibition studies with oligosaccharides of known structure also showed that the antibody binds specifically to glycoconjugates with an alpha 1----3 terminal galactose residue. Anti-Gal did not bind to a human B-active glycolipid, indicating that fucose-linked alpha 1----2 to the penultimate galactose prevents anti-Gal binding. The anti-Gal specificity for RRBC glycolipids also paralleled that of the alpha-galactosyl-specific Bandeiraea simplicifolia lectin. The possible reasons for the occurrence of this unique antibody in human serum are discussed.

Anti-trophoblast antibody responses during normal human pregnancy

During the course of a normal uncomplicated human pregnancy the mother generates an antibody response directed against determinants present on the plasma membrane of the outer fetal layer of the term placenta, the syncytiotrophoblast. The response, measured by an ELISA that utilises syncytiotrophoblast plasma membrane as the antigenic target, is predominantly IgG in nature, but with a minor contribution from IgM molecules. Maximum responses were observed during the first trimester and the levels gradually declined as the pregnancy progressed. On a population basis, this antibody response profile was mainly restricted to first and second pregnancies, although anti-trophoblast antibody responses could be detected in multiparous women but with a greatly reduced incidence compared with primipara. Mechanisms to account for these observations are discussed. Throughout, the anti-trophoblast antibody levels detected in pregnancy sera were compared with the background levels which were observed in sera obtained from males and nulliparous non-pregnant females.

A unique natural human IgG antibody with anti-alpha-galactosyl specificity.

A new natural anti-alpha-galactosyl IgG antibody (anti-Gal) was found to be present in high titer in the serum of every normal individual studied. The antibody was isolated by affinity chromatography on a melibiose-Sepharose column. The reactivity of the antibody was assessed by its interaction with alpha-galactosyl residues on rabbit erythrocytes (RabRBC). The specificity was determined by inhibition experiments with various carbohydrates. The anti-Gal interacts with alpha-galactosyl residues, possibly on glycolipids of human RBC (HuRBC), after removal of membrane proteins by treatment with pronase. In addition, the anti-Gal bind specifically to normal and pathologically senescent HuRBC, suggesting a physiological role for this natural antibody in the aging of RBC. The ubiquitous presence of anti-Gal in high titers throughout life implies a constant antigenic stimulation. In addition to the theoretical interest in the antibody, the study of the anti-Gal reactivity seems to bear immunodiagnostic significance. Decrease in the antibody titer was found to reflect humoral immunodeficiency disorders.

New synthetic and natural tuftsin-related compounds and evaluation of their phagocytosis-stimulating activity.

The preparation of large amounts of tuftsin (50 g) as well as purification by preparative HPLC are presented. Synthesis of several analogues exhibiting significant phagocytosis-stimulating activity of human PMN are described and discussed. On the other hand, the synthesis of a natural human IgG decapeptide fragment 335-344 that could be liberated from the same carrier molecule is presented. This decapeptide fragment, Thr-Ile-Ser-Lys-Ala-Lys-Gly-Gln-Pro-Arg, showed interesting activity in stimulating phagocytosis of human polymorphonuclear leukocytes.

Demonstration of a natural antigalactosyl IgG antibody on thalassemic red blood cells

A modified antiglobulin test, based on the high affinity between the Fc portion of the red blood cell (RBC) bound IgG and the Fc receptor on the myeloid cell K-562, was utilized for demonstration of immunoglobulins (Ig) on thalassemic RBC. Ig was found on the RBC of 73 out of 80 patients with thalassemia. The immunoglobulins on the thalassemic RBC belonged to the IgG subclass and were autoreactive. Elution studies utilizing various carbohydrates, or by thermal stripping, indicated that at least part of the IgG molecules found on the thalassemic RBC were specifically reactive with terminal galactosyl residues on the RBC membrane. IgG antibodies with similar reactivity were also demonstrated in normal human serum. These natural antigalactosyl IgG antibodies from normal sera could bind to IgG- depleted thalassemic RBC. Thalassemic RBC and normal senescent RBC were previously found to contain reduced amounts of membrane sialic acid (SA). It is suggested that the antigalactosyl IgG antibodies interact with newly exposed galactosyl residues underlying the sialic acid units. Such interaction may lead to the shortened lifespan of thalassemic RBC and may result in sequestration of senescent normal RBC by the reticuloendothelial system.

Demonstration of a Natural Antigalactosyl IgG Antibody on Thalassemic Red Blood Cells

A modified antiglobulin test, based on the high affinity between the Fc portion of the red blood cell (RBC) bound IgG and the Fc receptor on the myeloid cell K-562, was utilized for demonstration of immunoglobulins (Ig) on thalassemic RBC. Ig was found on the RBC of 73 out of 80 patients with thalassemia. The immunoglobulins on the thalassemic RBC belonged to the IgG subclass and were autoreactive. Elution studies utilizing various carbohydrates, or by thermal stripping, indicated that at least part of the IgG molecules found on the thalassemic RBC were specifically reactive with terminal galactosyl residues on the RBC membrane. IgG antibodies with similar reactivity were also demonstrated in normal human serum. These natural antigalactosyl IgG antibodies from normal sera could bind to IgG-depleted thalassemic RBC. Thalassemic RBC and normal senescent RBC were previously found to contain reduced amounts of membrane sialic acid (SA). It is suggested that the antigalactosyl IgG antibodies interact with newly exposed galactosyl residues underlying the sialic acid units. Such interaction may lead to the shortened lifespan of thalassemic RBC and may result in sequestration of senescent normal RBC by the reticuloendothelial system.

Multicomponent structure of the thyrotropin receptor: Relationship to Graves' disease

The thyrotropin receptor is proposed to contain both a glycoprotein and a ganglioside component. Monoclonal antibodies have been developed against soluble thyroid TSH receptor preparations and using Graves' lymphocytes. These show that initial recognition of thyrotropin requires the glycoprotein component, but that monoclonal antibodies to this component block thyrotropin function (“blocking antibodies”) rather than mimic thyrotropin. Monoclonal antibodies which stimulate thyroid activity in cultured cell systems (cAMP increase) or mouse bioassays, all interact with gangliosides. Using monoclonal antibodies to the glycoprotein component of the thyrotropin receptor, we show that two protein bands, molecular weights 18,000–23,000 and 50,000–55,000, are precipitated from detergent-solubilized preparations. Using a crosslinking procedure with 125I-labeled thyrotropin, we show that thyrotropin binding is related to the disappearance of the 18,000–23,000 molecular weight band on sodium dodecylsulfate gels and the appearance of a 30,000–33,000 molecular weight thyrotropin-membrane component complex. Higher molecular weight thyrotropin-membrane complexes of 75,000–80,000 and 250,000 are visualized when binding studies are performed at pH 7.4 in physiologic medium; larger amounts of the 30,000–33,000 complex are evident at pH 6.0 in a low salt medium. It is thus proposed that the glycoprotein component of the thyrotropin receptor is composed of two subunits with apparent molecular weights of 18,000–23,000 and 50,000–55,000; that the 18,000–23,000 subunit interacts with thyrotropin; and that different receptor subunits can exist depending on in vitro binding conditions. Using monoclonal-stimulating antibodies or natural autoimmune IgG preparations from patients' sera, we show that stimulating antibodies exhibit species-specific binding to human thyroid ganglioside preparations. The data unequivocally establish the pluritopic nature of the immunoglobulins in Graves' disease; demonstrate their effector potential in the pathogenesis of the disorder; and relate individual components or determinants of the thyrotropin receptor structure with specific autoimmune immunoglobulins.

The utility of human filarial serum in the detection of circulating antigen

The utility of human filarial serum immunoglobulin (FSI) in detecting circulating antigen in filarial sera was studied by counter immunoelectrophoresis (CIEP) and the indirect haemagglutination test (IHAT). CIEP was found to be better than IHAT. 23 out of 30 sera from persons with microfilaeremia and one of 30 clinical cases of filariasis, but none of the normal sera or sera from those with helminths other than filariae, showed the presence of circulating filarial antigen in CIEP. FSI was fractionated by DEAE- Sephadex A-50 column chromatography and the antibody active in CIEP was found to be IgG in nature.

Recognition of Heterologous Cells by Macrophages

Mouse and guinea pig macrophages cultured in vitro actively phagocytize non-opsonized thymocytes from certain heterologous animals including chickens, as shown in the accompanying paper (11). The present study was undertaken to investigate the mechanism of this phenomenon, using the phagocytosis of chicken thymocytes (c-thymocytes) by mouse and guinea pig macrophages. The involvement in this phenomenon of natural IgG passively adsorbed in situ to macrophages was excluded, since the phagocytosis of c-thymocytes was not significantly affected by the treatment of macrophages with homologous IgG or rabbit anti-sera directed toward homologous IgG. The involvement of lectin- or sugar-like receptors seems also to be unlikely, since various glycoproteins showed no significant effect. c-Thymocytes treated with normal mouse serum (NMS) but not with heat-inactivated NMS were strongly stained with goat anti-mouse C3 by an indirect immunofluorescent technique, and became extremely vulnerable to adherence to and phagocytosis by mouse macrophages, suggesting that c-thymocytes are an activator of the alternative pathway of mouse complement. These results as a whole raise the possibility that mouse and guinea pig macrophages can phagocytize c-thymocytes by recognizing their activating surfaces of the alternative complement pathway without the participation of exogenously added IgG or complement, as proposed by others in the phagocytosis of rabbit and mouse red blood cells by human monocytes.

Acquired von Willebrand’s Syndrome in the Course of Waldenström’s Disease

Acquired von Willebrand's syndrome with a regressive evolution is described in a 66 year old man with Waldenström's disease. An inhibitor electively directed against Ristocetin cofactor activity has been demonstrated, active in vitro after incubation at 37 degrees C. Serum fractionation showed that the inhibitor was independent of the monoclonal IgM and subsequent purification that it was IgG in nature. The results permit its classification as an auto-antibody.