The contribution of constant region domains to the binding of murine IgM to Fc mu receptors on T cells

IgM hybridoma constant region domain deletional mutants were used to investigate the domain requirements for binding of murine IgM to Fc mu receptors (Fc mu R) on normal murine T lymphocytes. Parental Sp 6:18 (mu, kappa; anti-trinitrophenyl) and its mutant proteins or their trinitrophenyl-antigen immune complexes were tested for their ability to inhibit the binding of pentameric IgM to Fc mu R on T lymphocytes. Inhibition was observed with ligands containing multiple copies of the third constant region domain. Inhibition did not occur with ligands missing the third constant region domain. In addition, a battery of rat monoclonal antibodies specific for individual murine IgM constant region domains was tested for the ability to inhibit the binding of pentameric murine IgM to Fc mu R on normal murine T lymphocytes. Total inhibition was observed with the antibodies directed to different epitopes located in C mu 3, but significant inhibition was not observed with antibodies directed to C mu 1, C mu 2, or C mu 4. Studies with domain deletional mutants and anti-domain antibodies have independently provided strong evidence that the C mu 3 domain plays a major role in the binding of IgM to Fc mu R on T lymphocytes and that C mu 1, C mu 2, and C mu 4 are not essential for binding. These studies have also provided evidence that valency and avidity influence the binding of IgM to T lymphocytes that express Fc mu R.     

Enhanced HIV-1 neutralization by a CD4-VH3-IgG1 fusion protein

HIV-1 gp120 is an alleged B cell superantigen, binding certain VH3+ human antibodies. We reasoned that a CD4-VH3 fusion protein could possess higher affinity for gp120 and improved HIV-1 inhibitory capacity. To test this we produced several human IgG1 immunoligands harboring VH3. Unlike VH3-IgG1 or VH3-CD4-IgG1, CD4-VH3-IgG1 bound gp120 considerably stronger than CD4-IgG1. CD4-VH3-IgG1 exhibited ≈1.5-2.5-fold increase in neutralization of two T-cell laboratory-adapted strains when compared to CD4-IgG1. CD4-VH3-IgG1 improved neutralization of 7/10 clade B primary isolates or pseudoviruses, exceeding 20-fold for JR-FL and 13-fold for Ba-L. It enhanced neutralization of 4/8 clade C viruses, and had negligible effect on 1/4 clade A pseudoviruses. We attribute this improvement to possible pairing of VH3 with CD4 D1 and stabilization of an Ig Fv-like structure, rather than to superantigen interactions. These novel findings support the current notion that CD4 fusion proteins can act as better HIV-1 entry inhibitors with potential clinical implications.     

Dextran sulfate and heparin interact with CD4 molecules to inhibit the binding of coat protein (gp120) of HIV

Dextran sulfate, heparin, and certain other sulfated polysaccharides potently inhibit the adsorption of HIV to CD4+ cells. The mechanism of this inhibition is unclear and, specifically, it is unknown if these agents act at the level of CD4-gp120 binding. For example, previous reports have demonstrated that dextran sulfate does not inhibit the cell surface binding of anti-CD4 mAb known to be directed at the gp120 binding site. In order to confirm and extend these observations, in the present study, it was shown that dextran sulfate does not inhibit the binding of OKT4A, OKT4C, Leu3a, or B66.6 to CD4+ cells as measured by cytofluorography. Next, recombinant forms of CD4 (rT4) and gp120 (rgp120) were utilized to directly study their molecular interaction in the absence of other viral or cellular structures. Reciprocal solid phase ELISA assays were developed to study directly the effects of sulfated polysaccharides on the binding of rT4 to immobilized rgp120 and vice versa. Dextran sulfate, heparin, and fucoidan, but not chondroitin sulfate, inhibited the binding of rgp120 to rT4. Importantly, dextran sulfate and heparin pre-treatment of immobilized rT4, but not immobilized rgp120, inhibited rT4-rgp120 binding. Taken together, these data suggest that while both sulfated polysaccharides and anti-CD4 mAb inhibit gp120 binding, the sulfated polysaccharides interact with sites on CD4 that are distinct from those with which the antibodies bind.     

Dichotomous effect of monocyte Fc receptor interaction on anti-CD3-induced immunoglobulin synthesis

Monoclonal antibodies against the TCR/CD3 complex are capable of activating T cells which in turn may induce immunoglobulin synthesis in B cells under appropriate conditions. Here we present evidence that distinct immune responses, induced by four commonly used TCR/CD3 mAb (Leu4, OKT3, BMA030, BMA031) were related to the mAb interaction with monocyte Fc receptors for IgG. Depending on their isotype and on the technique by which they were crosslinked, TCR/CD3 mAb induced variable IgM and IgG synthesis in PBMC: If the mAb were crosslinked by monocyte IgG-Fc receptors they induced a high Ig production, while crosslinking the same mAb by plastic-bound goat anti-mouse antibodies (panning) failed to do so. Nevertheless, both crosslinking techniques triggered a strong proliferation and IL-2, IL-4, and IFN gamma lymphokine gene expression. The lack of Ig production under panning conditions was due to an additional IgG-Fc receptor interaction with monocytes: (a) If namely mAb F(ab')2 fragments, or mAb isotypes unable to bind to monocyte Fc receptors (IgG2b, IgG1 in nonresponders) were crosslinked by panning, both a good proliferation as well as Ig production ensued; (b) if TCR/CD3 mAb isotypes which could additionally bind to monocyte Fc receptor (IgG2a) were crosslinked, no Ig production occurred; (c) if mAb F(ab')2 fragments were crosslinked with a second anti-T cell antibody of IgG2a isotype, which could bind to monocyte Fc receptors, Ig synthesis was reduced. Interestingly enough, this diminishing effect, due to monocyte Fc receptor interaction, was only observed if CD4-positive cells were proliferating, but not if CD8-positive cells were activated.     

Fc receptors and immunoglobulin binding factors

Receptors for the Fc portion of Ig (Fc receptors, FcR) are found on all cell types of the immune system. Three types of FcR react with IgG: Fc gamma RI is a high-affinity receptor binding IgG monomers whereas Fc gamma RII and Fc gamma RIII are low-affinity receptors binding IgG immune complexes; the three types of Fc gamma R are members of the Ig superfamily. Two FcR react with IgE:Fc epsilon RI is a multichain receptor binding IgE with high affinity; it is composed of an IgE-binding alpha chain, homologous to Fc gamma RIII, and of gamma and beta chains that are necessary for receptor expression and signal transduction. The low-affinity Fc epsilon RII is the only FcR described so far that is not a member of the Ig superfamily but resembles animal lectins; it is composed of a transmembrane chain with an intracytoplasmic NH2 terminus. Fc alpha R has homology with Fc gamma R and is a member of the Ig superfamily. Receptors for IgM and IgD are not characterized yet. Finally, Ig transport is made by FcR-like molecules such as the poly-Ig receptor or an MHC-like receptor found on neonatal intestine. A remarkable property of most FcR is the fact that they are released in cell supernatants and circulate in biological fluids as immunoglobulin binding factors (IBF) generated either by cleavage at the cell membrane or by splicing of FcR transmembrane exon. Immunoglobulin binding factors may interfere with Ig-mediated functions and have direct immunoregulatory activities. Involvement of FcR or IBF has been postulated in several diseases, and monoclonal antibodies to FcR are beginning to be used in therapeutics, particularly to target cytotoxic effector lymphocytes and monocytes to tumor cells.     

Mouse Ig coded by VH families S107 or J606 bind to protein A

Twenty-five monoclonal mouse Ig (5 IgA, 7 IgM, and 13 IgG1) were tested for binding to staphylococcal protein A. They were allowed to attach to protein A Sepharose column at pH 8.0 and were then eluted with a pH gradient from approximately 7.5 to 3.0. Five of them (IgM or IgA) did not bind. Ten came off with pH approximately 6. They were all IgG1, and were probably bound (weakly) via the Fc portion. The remaining 10 (3 IgA, 4 IgM, and 3 IgG1) were more firmly bound; they came off with pH-values ranging from 5.0 to 3.5. They all expressed VH genes of families J606 or S107, whereas all the 15 Ig that were not firmly bound expressed VH genes of six other families. The VL domains seem to be unimportant for protein A binding inasmuch as a firmly binding and a weakly binding IgG1 antibody share identical VK domains. VH sequences of protein A-binding and nonbinding Ig were compared. No likely peptide sequences were found that might make the ligand for protein A.     

IL-4 (B cell stimulatory factor 1) overcomes Fc gamma receptor-mediated inhibition of mouse B lymphocyte proliferation without affecting inhibition of c-myc mRNA induction

Mouse B cells are stimulated to proliferate by Fab'2 fragments of rabbit anti-mouse Ig antibodies. Proliferation is inhibited, however, in the presence of IgG anti-mouse Ig. We have previously shown that this inhibition is mediated by binding of the IgG anti-Ig to receptors for Fc gamma R on B cells. This report describes conditions under which IgG anti-mu or anti-delta will induce proliferation despite Fc gamma R engagement. Culture supernatants of Con A-stimulated, Il-4-secreting Th cell lines, but not of Il-2-secreting Th cell lines, will co-stimulate with IgG anti-Ig to induce small B cells to incorporate [3H]TdR. This co-mitogenic activity is inhibitable by anti-IL-4 antibodies and can also be induced by Il-4 affinity purified from the T cell supernatants or by supernatants containing rIl-4. B cells precultured with Il-4 for 18 h, while still expressing normal levels of Fc gamma R, also proliferate to IgG anti-Ig. We have previously shown that Fc gamma R-mIg cross-linking will inhibit mIg-dependent increases in c-myc mRNA levels. We investigated whether Il-4 allows B cells to respond to IgG anti-Ig by elevating c-myc. The data show that Il-4 has little effect on c-myc mRNA levels in either IgG or Fab'2 anti-Ig-containing cultures.     

Structural characterization of a recombinant CD4-IgG hybrid molecule

CD4-IgG is a homodimer of a hybrid polypeptide consisting of the two amino-terminal domains (residues 1-180) of human CD4 fused to the hinge region and the second and third constant-sequence (CH2 and CH3) Fc domains (residues 216-441) of human immunoglobulin G (IgG-1). This antibody-like molecule, termed an immunoadhesin, was produced in an effort to combine the binding specificity of CD4 with several potentially desirable properties of IgG molecules [Capon et al. (1989) Nature 337, 525-531]. The structural characteristics of the molecule have been evaluated to demonstrate that CD4-IgG has the same features as the N-terminal region of soluble CD4, while retaining those expected for the Fc portion of human IgG. Identification of peptides recovered from the tryptic map confirmed 98.8% of the expected structure of CD4-IgG. The detection of glucosamine in peptides containing Asn257 and the retention time shift of this tryptic peptide after deglycosylation confirmed the presence of Asn-linked oligosaccharides at this position. Four pairs of intrachain and two interchain disulfide bonds were also established.     

Erythrocyte membrane-associated immunoglobulins during malaria infection of mice.

Immunoglobulin (Ig) is associated with erythrocyte membranes during infection of A/J mice with Plasmodium berghei. It was detected by agglutination of the cells with rabbit antibodies to mouse IgG and by a radioimmunoassay with 125I-labelled rabbit antibodies to mouse IgG. As shown by the degree of agglutination and of binding of radiolabeled antibodies to the erythrocyte membranes, the amount of Ig increased with time after infection and paralleled parasitemia and reticulocytosis. The erythrocyte-associated immunoglobulins are mainly IgG but IgM was also present on the cells of some mice. A large proportion of the Ig could be eluted at 37 degrees C and was analyzed by immunoprecipitation and acrylamide gel electrophoresis. A radioautographic study with 125I-labeled anti-mouse IgG revealed that both parasitized and nonparasitized reticulocytes of infected mice had much larger amounts of membrane-bound immunoglobulin than did mature, nonparasitized erythrocytes. The nature of the bonds between the Ig and the surface membrane of the reticulocytes is not known. The Ig could be part of immune complexes nonspecifically bound to the cell surface. However, since we have not detected Fc or C3d receptors on reticulocytes, it is possible that the Ig constitutes autoantibodies against reticulocytes or antibodies against parasitic antigens present on the cell membrane.     

Antigen-antibody complexes bound to B-lymphocyte Fc gamma receptors regulate B-lymphocyte differentiation

We studied the effect of soluble antigen-antibody complexes on the responses of polyclonally activated murine B lymphocytes. For this, normal B lymphocytes were stimulated with rabbit F(ab')2 anti-mu and lymphokines. IgG complexes, particularly in antigen excess, inhibited the plaque-forming cell response (55-70%), while proliferation was unaffected. Maximal inhibition was obtained with small amounts (0.2-1.0 microgram/ml) of complexes. Neither antigen or antibody alone was inhibitory. Inhibition was mediated via binding of the IgG complexes to Fc gamma receptors of B lymphocytes: (1) neither T lymphocytes or adherent accessory cells were required; (2) IgM complexes did not inhibit; and (3) inhibition was not seen when monoclonal anti-Fc gamma receptor antibodies prevented binding of the IgG complexes to these receptors. Kinetic experiments showed that B lymphocytes are susceptible to this inhibitory signal for only a short time after stimulation. We conclude that IgG complexes bound to the Fc gamma receptors of B lymphocytes regulate B-lymphocyte differentiation.     

Leukotriene B4 potentiates the expression and release of Fc epsilon RII/CD23, and proliferation and differentiation of human B lymphocytes induced by IL-4

This study documents the influence of leukotriene (LT) B4 on human B lymphocyte responses. Incubation of freshly isolated B lymphocytes with LTB4, but not LTC4, induced a slight but significant, time- and dose-dependent increase in the surface expression of Fc epsilon RII/CD23 and class II MHC Ag and in the release of soluble CD23. These changes were maximal at 10 nM LTB4 after an incubation period of 48 h. When B lymphocytes were preactivated in vitro with Staphylococcus aureus Cowan strain I (SAC), neither LTB4 nor LTC4 was able to promote proliferation and/or IgG and IgM secretion. In contrast, when resting B lymphocytes were stimulated with a suboptimal concentration (3 U/ml) of IL-4, LTB4, but not LTC4, potentiated both the Fc epsilon RII/CD23 and the class II MHC antigen expression, and the release of soluble CD23 in a dose-dependent manner, without affecting the kinetics of these responses. Furthermore, LTB4, but not LTC4, amplified both the proliferative response and the IgG and IgM secretion induced by addition of a suboptimal dose of IL-4 (3 U/ml) to SAC-preactivated B lymphocytes. Again, LTB4 did not modify the kinetics of the proliferative response promoted by IL-4. Although LTB4 potentiated IL-4-induced IgG and IgM secretion from SAC-activated B lymphocytes, no production of IgE was observed. These data indicate that LTB4 could play a regulatory role in the modulation of IL-4-induced signaling in human B lymphocytes.     

Human IgA and IgG F(ab')2 that bind to staphylococcal protein A belong to the VHIII subgroup

Staphylococcal protein A (SPA) is a bacterial membrane protein that possesses, in addition to its Fc gamma-binding activity, a distinct specificity for the Fab region of some IgM, IgA, IgG, and IgE. The Fab site that binds to SPA has been localized to the V region of the Ig H chain. In a previous study of human monoclonal and polyclonal IgM, we demonstrated that binding to SPA was highly restricted to molecules of the VHIII subgroup, and that nearly all VHIII IgM were able to bind SPA. The present study examines the VH composition of SPA-binding and SPA-nonbinding fractions of purified human polyclonal IgA, and IgG F(ab')2 fragments. We found that 22% of the IgA and 15% of the IgG F(ab')2 bound to SPA-agarose. Analysis with VH subgroup-specific antisera indicated that the SPA-binding fraction of IgA was dominated by the VHIII subgroup, and the SPA-binding fraction of IgG F(ab')2 contained only VHIII molecules. Furthermore, substantial portions of the total VHIII protein in IgA and in IgG F(ab')2 bound to SPA. We conclude that Fab binding to SPA is both restricted to and highly prevalent among human VHIII molecules, regardless of Ig class. These results suggest that protein A is an Ig superantigen.     

Trypanosoma musculi co-express several receptors binding rodent IgM, IgE, and IgG subclasses

The present work demonstrates the expression of receptors for the Fc portion of rodent Ig by the murine parasite Trypanosoma musculi. By using a rosette assay adapted to the parasite morphology and by flow cytometry analysis, three distinct receptors were identified. A receptor binding rabbit or rat polyclonal IgG and mouse monoclonal IgG1, IgG2a, and IgG2b was found on parasites purified from the blood and the peritoneal cavity of infected mice and on parasites maintained in culture conditions. This IgG receptor was degraded by pepsin. A separate receptor, binding only mouse monoclonal IgG3 was observed on cultured parasites. A receptor binding rabbit, rat, and mouse IgM was found on cultured and peritoneal parasites, but not on blood parasites. This receptor did not bind IgG or IgA but it bound mouse and rat IgE as well as IgM. It was degraded by trypsin. IgG and IgM/IgE receptors were co-expressed on single parasites. They were not of host origin but synthesized by trypanosomes as shown by reexpression in vitro after proteolytic degradation. Their expression was variable with the development of trypanosomes both in vitro and in vivo.     

人IL35-IgG4(Fc)融合蛋白在CHO/DG44细胞中的稳定表达

本研究利用基因重组技术构建人IL35-IgG4(Fc)融合基因真核表达载体, 稳定转染CHO/DG44细胞并检测重组蛋白的表达。主要采用聚合酶链式反应(PCR)从脂多糖(Lipopolysaccharides, LPS)诱导的人髓性白血病细胞株KG-I cDNA文库中克隆EBI3和IL-12p35 cDNA, 重叠PCR法连接2个片段, 并克隆到IgG4(Fc)- pOptiVEC?-TOPO?载体上,对新构建的IL-35-IgG4 (Fc) pOptiVEC?-TOPO?真核表达载体并进行酶切、测序、PCR鉴定; 脂质体法转染CHO/DG44细胞; RT-PCR检测转染结果, 采用a-MEM-培养基筛选实验组细胞, 对筛选的阳性克隆细胞再进行氨甲喋呤(Methotrexate, MTX)的加压筛选, ProteinG-Agarose纯化阳性克隆培养上清, 免疫印迹检测目的蛋白表达。结果显示IL-35-IgG4 (Fc) pOptiVEC?-TOPO?表达载体稳定转染CHO/DG44细胞并获得阳性克隆; SDS-PAGE电泳得到一条与预期相对分子质量大小相符的蛋白条带; 该蛋白能与羊抗人IgG4抗体特异结合。本实验获得了能够稳定表达具有稳定结构的IL35-IgG4(Fc)融合蛋白的CHO/DG44细胞株。     

Bi-specific monoclonal antibodies: selective binding and complement fixation to cells that express two different surface antigens

A new dimension of immunotherapeutic selectivity might be achieved if antibodies could distinguish cells that co-express two different surface antigens. Bi-specific monoclonal antibodies (BSMAB) with two different antigen combining sites that share a common Fc region theoretically might have such a potential. Two such BSMAB were produced by hybrid-hybridoma clones prepared by fusion of pre-existing hybridomas and were purified by isoelectric focusing. CD3,4 (IgG2a, IgG2b) recognizes the T cell surface antigens CD3 and CD4, and CD3,8 (IgG2a, IgG2a) recognizes CD3 and CD8. These BSMAB promote complement-mediated lysis of target cells that bear both surface antigens 25 to 3125 times more efficiently than those that express only one of the antigens. This selectivity results from the increased avidity of these antibodies for cells with both antigens, as reflected by the increased surface immunoglobulin concentration detected by flow cytometry. It was also demonstrated that there exists a threshold surface immunoglobulin density necessary for antibody-dependent complement-mediated cytotoxicity microtiter assays for the various IgG antibodies tested in both bivalent and monovalent binding. These results support the associative model of IgG-mediated complement fixation.     

Monoclonal antibodies to Fc receptors for IgG on human mononuclear phagocytes. Antibody characterization and induction of superoxide production in a monocyte cell line

We have utilized monoclonal antibodies against the two IgG Fc receptors (p40 and p72) of U937 cells to stimulate the release of superoxide. The monoclonal antibody (mAb) specific for p40 (IV3) has been described elsewhere. A murine IgG1 mAb specific for the high affinity p72 Fc receptor (designated mAb FcR32 or simply mAb 32) bound to the same p72 precipitated by Sepharose-human IgG as shown by preclearing experiments and by identical isoelectric focussing patterns. Binding of mAb 32 to p72 was independent of the Fc region of the antibody since Fab' fragments of mAb 32 affinity adsorbed p72. The binding of both mAb 32 and human IgG1 to the intact U937 cell was not reciprocally inhibitory, indicating that mAb 32 does not interfere with the ligand binding site of p72. mAb 32 bound to human monocytes, U937, and HL60 cells, but not to granulocytes or lymphocytes. U937 cells cultured in gamma-interferon and 1,25-dihydroxycholecalciferol generated superoxide when incubated with mAb 32 or IV3 followed by cross-linking with F(ab')2 anti-murine Ig. Incubation with mAb 32 or IV3 alone or with 3 of 5 other anti-U937 mAbs cross-linked with anti-murine Ig did not result in superoxide generation. Immune complex-mediated superoxide production was inhibited 80% by IgG, but not by mAb 32 or IV3.     

Interference of rheumatoid factor activity by aspartame, a dipeptide methyl ester

Circulating autoimmune complexes of IgM rheumatoid factors (RF) bound to the Fc portions of normal, polyclonal IgG antibodies are frequently present in humans with rheumatoid arthritis (RA). The sweet tasting methyl ester of L-Asp-L-Phe (aspartame or APM) was found to relieve pain and improve joint mobility in subjects with osteo- and mixed osteo/rheumatoid arthritis [Edmundson, A. B. and Manion, C. V. (1998). Clin. Pharmac. Ther. 63, 580-593]. These clinical observations prompted the testing of the inhibition by APM of the binding interactions of human IgM RFs with IgG Fc regions. The propensity of APM to inhibit IgM RF binding was assessed by competitive enzyme immunoassays with solid-phase human IgG. Ten RA serum samples and three purified monoclonal cryoglobulins, all of which had RF activity, were tested in this system. We found that the presence of APM significantly reduced the binding of IgM RFs. The inhibitory propensity of APM with monoclonal RF cryoglobulins was increased by the addition of CaCl(2) to the binding buffer. Similar inhibition of the binding of RA derived RFs to IgG was observed for Asp-Phe and its amidated derivative, indicating that the methyl ester is not required for APM's interaction with IgM antibodies. A human (Mez) IgM known to bind octameric peptides derived from the Fc portion of a human IgG(1) antibody was tested for binding of dipeptides by the Pepscan method of combinatorial chemistry. The relative binding constants of Asp-Phe and Phe-Asp were ranked among the highest values for 400 possible combinations of the 20 most common amino acids. Possible blocking interactions of APM were explored by computer-assisted docking studies with the model of a complex of an RF Fab with the Fc of a human IgG(4) antibody. Modeling of ternary immune complexes revealed a few key residues, which could act as molecular recognition sites for APM. A structural hypothesis is presented to explain the observed interference with RF reactivity by APM. Extrapolations of the current results suggest that APM may inhibit the binding of IgG in a substantial proportion of IgM RFs. Interference of RF reactivity, especially in RA patients, may alleviate the pain and immobility resulting from chronic inflammation of the joints.     

IgM expressed by leukemic CD5(+) B cells binds mouse immunoglobulin light chain

Mouse immunoglobulin (Ig) molecules have previously been shown to bind to the surface of CD5(+) B cells from patients with B-cell chronic lymphocytic leukemia (B-CLL). The results indicated that surface IgM was involved in the interaction and suggested the phenomenon was an example of the polyreactive binding capacity of the surface Ig (sIg) expressed by these malignant cells. This article describes the further characterization of the interaction between human IgM and mouse Ig molecules and subunits. Mouse Ig molecules of both kappa and lambda light chain classes bound to the B-CLL cell surface. The dissociation constant for the interaction of mouse IgG1 (K121) with the B-CLL cell surface was 3.6 x 10(-7) M. To confirm the involvement of the human IgM expressed by the B-CLL cells in the interaction, the malignant cells were stimulated in vitro to induce secretion of human IgM. Enzyme immunoassay was used to show that secreted human IgM bound to intact mouse Ig, as occurred with the cell surface analysis. The mouse Ig epitope recognized by the purified secreted human IgM was shown by Western blot analysis to be located on the light chain of the mouse Ig molecule and to be conformationally dependent. K121 light chain was cloned and expressed in E. coli and the recombinant light chain bound to the surface of CLL B cells. The results confirm that human IgM is the reactive ligand in the interaction with mouse Ig and indicate that the interaction of polyreactive IgM with mouse IgG occurs via the light chain component of IgG.     

Bindings of plasma proteins to streptococci of serological group L with special reference to their immunoglobulin G Fc-receptor activity

Of 33 streptococcal cultures belonging to serological group L, all bound human immunoglobulin (Ig) G, fibrinogen, and fibronectin; 32 bound bovine IgG; 31 bound alpha 2-macroglobulin; 5 bound albumin; and none bound either haptoglobin or IgA. The binding sites for IgG could be isolated from the L streptococci by trypsinization and purified by affinity chromatography on human IgG-Sepharose. The purified Fc receptors reacted with IgG subclasses 1, 2, 3, 4 of humans, 1 and 2 of bovines, ovines, and caprines as well as a, b, c, and T of equines. They had a molecular mass of approximately 49,000 Da. Thus, the Fc receptors from L streptococci corresponded to type III Fc receptors of Streptococcus dysgalactiae.     

An appraisal of Fc receptors on human peripheral blood B and L lymphocytes.

Human circulating lymphocytes with easily detectable surface immunoglobulin have been divided into two populations, B cells and L cells. This second population lacks membrane-incorporated Ig, but has a receptor for membrane-labile cytophilic IgG. In this study purified B and L lymphocytes were examined for Fc receptors that bind aggregated IgG and IgG complexed to erythrocytes. Purified lymphocyte populations were prepared by nylon columns and by negative selection with rosetting techniques. L lymphocytes bound aggregated guinea pig and human IgG, and formed rosettes with human erythrocytes sensitized with Ripley IgG (EA). Treatment of L lymphocytes with trypsin had no effect on the receptors for IgG. B lymphocytes did not bind EA and attachment of aggregated IgG was variable; up to one-third of these cells fixed aggregated human IgG to the cell membrane. Trypsin treatment abolished binding of Agg-IgG to B cells in sharp contrast to its effect on L cells. Furthermore, double-label immunofluorescence studies showed that cells with both membrane-incorporated Ig and receptors for aggregated guinea pig IgG were rare. These studies indicate that human peripheral blood B lymphocytes lack a high affinity, trypsin-resistant Fc receptor that is present on L lymphocytes.