Different rosette assays for detecting Fc receptor-bearing lymphocytes measure different subpopulations

The capacity of purified lymphocytes from human peripheral blood to bind the Fc portion of IgG was investigated by the rosette technique using ox erythrocytes sensitized with rabbit anti-ox IgG (EA_ox) and human erythrocytes sensitized with anti-CD IgG (EA_CD). With unfractionated lymphocytes EA_ox always detected more rosette-forming cells (RFC) than did EA_CD; however, in lymphocyte populations specifically depleted of B lymphocytes by passage through copolymer styrene bead columns, the proportion of rosettes formed with EA_ox and with EA_CD was the same. Double labelling for Fc receptors and surface immunoglobulin (SIg) demonstrated that most of the lymphocytes which formed rosettes with either EA_ox or EA_CD also carried SIg. Pre-incubation of lymphocytes at 37° to remove heatlabile SIg did not affect their ability to form EA rosettes but reduced the proportion of SIg-bearing cells. Following pre-incubation a significant proportion of EA_ox RFC still remained SIg positive but the lymphocytes which formed rosettes with EA_CD no longer carried SIg.

These studies suggest that rosette formation with EA_CD detects only `K' lymphocytes (non-T, non-B cells bearing heat-labile SIg) while EA<sub>ox</sub> detects some B lymphocytes as well. By reducing the amount of IgG bound to ox erythrocytes sensitivity was reduced to the point where EA<sub>ox</sub> no longer formed rosettes with B lymphocytes but still detected `K' lymphocytes indicating either a qualitative or quantitative difference between the Fc receptors on B and `K' lymphocytes. Treatment of lymphocytes with trypsin decreased the percentage of rosettes formed with EA<sub>ox</sub> but not with EA<sub>CD</sub> supporting the conclusion that there is a structural difference between the Fc receptors on B and `K' lymphocytes although a difference in receptor density is not excluded.

When EA_CD and fluorescein-labelled ox erythrocytes sensitized with a low concentration of rabbit anti-ox IgG were mixed, most RFC bound both the indicator erythrocytes simultaneously suggesting that the Fc receptors on `K' lymphocytes do not exhibit species specificity.

     

Influence of IgG and IgM receptor triggering on human natural killer cell cytotoxicity measured on the level of the single effector cell

By using an agarose single cell cytotoxicity assay, in combination with rosetting with IgG- or IgM-coated ox red blood cells for detection of Fc receptors for IgG (FcγR) or IgM (FcμR), it was found that 60% of natural killer (NK) cells are FcγR⁺ and 17% FcμR⁺. This system was further used to investigate the consequence of FcμR and FcγR triggering on NK killing as measured on the single effector cell level. It was found that stimulation of the FcγR, but not the FcμR, resulted in substantial NK inhibition. In order for NK cells to be inhibited by IgG-coated ox red blood cells, they must first be exposed to the IgG-containing complex prior to conjugation with the target. While exposure of the FcμR to immune complexes can block up to 57% of NK activity, the particulate immune complexes do not interfere with binding of effector cells to targets. Modulation of Fcγ R by capping at 37°C does not interfere with the NK inhibition for up to 3 h, though after 20 h, when FcγR⁺ cells are almost completely modulated, NK activity has fully returned. Although to a lesser extent, the soluble immune complex human transferrin-anti-transferrin also reduces NK cell activity when activating effector cell FcγR prior to target cell binding. Also, pretreatment of target cells with high concentrations of specific antibody toward membrane antigens can block NK activity while not inhibiting target cell binding as evidenced by anti-IgM IgG binding to Daudi cells. The regulatory influence of the FcγR on the NK system is discussed in terms of other functions associated with this receptor and in terms of its possible biological significance.     

Affinity of the interaction between Fcgamma receptor type III (FcγRIII) and monomeric human IgG subclasses. Role of FcγRIII glycosylation

Binding of the Fc region of IgG antibodies to low affinity Fcγ receptors (FcγR) triggers important effector functions in the immune system. The type IIIb FcγR (FcγRIIIb or CD16) is a heavily glycosylated protein anchored to the membrane of neutrophils by a glycosylphosphatidylinositol link. This receptor contributes to cell activation by IgG immune complexes. To better understand the nature of the ligand-receptor association, we have studied the affinity and kinetics of the interaction between human IgG subclasses and two soluble forms of FcγRIIIb (sFcγRIIIb or sCD16) corresponding to the 188 N-terminal amino acids of the extracellular region of the receptor, a glycosylated one made in eucaryotic cells (euc.sCD16) and a non-glycosylated one (proc.sCD16) made in Escherichia coli. Experiments using a BIAcore^TM instrument, to measure protein binding in real time, showed that monomeric human IgG1 and IgG3, but not IgG2, IgG4, IgA and divalent antigen-binding fragments (F(ab′)₂) of IgG1, bound to immobilized euc.sCD16 with an affinity constant (K_A) of 1.3 ± 0.6 × 10⁶ M^?1 and 2.6 ± 0.4 × 10⁵ M^?1, respectively. The affinity constant of proc.sCD16 for human IgG1 was in the same range (1.1 ± 0.2 × 10⁶ M^?1), whereas that for human IgG3 was twofold higher (4.2 ± 0.4 × 10⁵ M^?1). The specificity of the non-glycosylated receptor for human IgG subclasses bound to Sepharose was IgG1 > IgG3 ? IgG4 ? IgG2. Thus, the extracellular polypeptide of FcγRIIIb dictates the interaction of the receptor with IgG subclasses although glycosylation plays an inhibitory role in the interaction with human IgG3.     

Both FcγRIV and FcγRIII are essential receptors mediating type II and type III autoimmune responses via FcRγ‐LAT‐dependent generation of C5a

FcγRIV is a relatively new IgG Fc receptor (FcγR) that is reported to contribute to the pathogenesis of autoimmune diseases, although its specific role in relation to FcγRIII, complement and IgG2 subclasses remains uncertain. Here we define FcγRIV on macrophages as a receptor for soluble IgG2a/b complexes but not for cellular bound IgG2a and show that simultaneous activation of FcγRIV and FcγRIII is critical to mediate certain type II/III autoimmune responses. FcγRIII‐deficient mice display compensatory enhanced FcγRIV expression, are protected from lung inflammation after deposition of IgG complexes, and show reduced sensitivity to IgG2a/b‐mediated hemolytic anemia, indicating that increased FcγRIV alone is not sufficient to trigger these diseases in the absence of FcγRIII. Importantly, however, blockade of FcγRIV is also effective in inhibiting phagocytosis and cytokine production in IgG2b‐induced anemia and acute lung injury, processes that display a further dependence on C5a anaphylatoxin receptor. Using gene deletion and functional inhibition studies, we found that FcγRIII and FcγRIV are each essential to trigger an FcRγ‐linker for activation of T‐cell‐dependent signal that drives C5a production in the Arthus reaction. Together, the results demonstrate a combined requirement for FcγRIII and FcγRIV in autoimmune injury, and identify the linker for activation of T cells adaptor as an integral component of linked FcγR and C5a anaphylatoxin receptor activation to generate inflammation.     

Dengue virus neutralization is modulated by IgG antibody subclass and Fcγ receptor subtype

Severe dengue virus (DENV) infection is epidemiologically linked to pre-existing anti-DENV antibodies acquired by maternal transfer or primary infection. A possible explanation is that DENV immune complexes evade neutralization by engaging Fcγ receptors (FcγR) on monocytes, natural targets for DENV in humans. Using epitope-matched humanized monoclonal antibodies (mAbs) and stable FcγR-transfected CV-1 cells, we found that DENV neutralization by IgG1, IgG3, and IgG4 mAbs was enhanced in high-affinity FcγRIA transfectants and diminished in low-affinity FcγRIIA transfectants, whereas neutralization by IgG2 mAbs (low-affinity ligands for both FcγRs) was diminished equally. In FcγR-negative Vero cells, IgG3 mAbs exhibited the strongest neutralizing activity and IgG2, the weakest. Our results demonstrate that DENV neutralization is modulated by the Fc region in an IgG subclass manner, likely through effects on virion and FcγR binding. Thus, the IgG antibody subclass profile generated by DENV infection or vaccination may independently influence the magnitude of the neutralizing response.     

Stimulation of the intracellular killing of Staphylococcus aureus by human monocytes mediated by Fcγ receptors I and II

Previous studies have shown that intracellular killing of bacteria by monocytes is stimulated by interaction between IgG and Fey receptors (FcγR) in the membrane of these cells. In the present study anti-FcγR monoclonal antibodies (mAb) were used to investigate the relative contributions of the various classes of FcγR to the intracellular killing of Staphylococcus aureus by human monocytes and the biochemical pathways involved. Anti-FcγRI or anti-FcγRII mAb, but not anti-FcγRIII mAb, efficiently stimulated the intracellular killing of bacteria by monocytes. Cross-linking FcγRI or FcγRII, but not FcγRIII, on monocytes with mouse anti-FcγR mAb followed by bridging with F(ab′)₂ fragments of goat anti-mouse IgG enhanced this process. Since the NADPH oxidase inhibitor diphenyleneiodonium blocked the FcγR-mediated intracellular killing of S. aureus, oxygen-dependent bactericidal mechanisms are most probably involved. Cross-linking FcγRI or FcγRII but not binding of the mAb to the FcγR on monocytes activated phospholipase C, as demonstrated by the increase in the intracellular concentration of inositol- (1,4,5)-triphosphate. The enhanced intracellular killing stimulated by cross-linking FcγR on monocytes was completely blocked by U-73122, an inhibitor of phospholipase C-dependent processes. Protein kinase C activity, but not the rise in the cytosolic free Ca⁺⁺ concentration or pertussis toxin-sensitive G proteins, is essential for the FcγR-mediated intracellular killing of bacteria by monocytes. Together, these results demonstrate that cross-linking FcγRI or FcγRII is equally effective in stimulating the intracellular killing of bacteria by monocytes and that this stimulation is a phospholipase C-dependent process.     

Quantitative contributions of IgG,IgM and C3 to erythrophagocytosisand rosette formation by peritoneal macrophages,and anti-opsoninactivity of dextran sulfate 500

In vitro phagocytosis by guinea pig peritoneal macrophages of immune complexes (EA) was shown to be dependent on IgG antibody in a dosedependent fashion. C3b enhanced phagocytosis of EA at limited IgG antibody concentrations only. When IgM antibody was used for sensitzation ofsheep red blood cells (SRBC), phagocytosis and rosette formation did notoccur in the absence of bound C3. The polyanion, dextran sulfate 500 (DS), was shown to depress both rosette formation and phagocytosis of EA_IgG, EA_IgGC1423 and EA_IgMC1423, as well as immune adherence of human group 0 erythrocytes and hemolytic activity of C3. This effect of DS was seen only when it was actually present in the incubation medium.     

FcγRIa–γ-chain complexes trigger antibody-dependent cell-mediated cytotoxicity (ADCC) in CD5+ B cell/macrophage IIA1.6 cells

Most receptors for immunoglobulins exist as multi-subunit complexes, with unique ligand binding α-chains, combined with accessory signalling (γ-, β-, or ζ-) chains. The myeloid class I receptor for IgG (FcγRIa) has been shown to be dependent on the FcR γ-chain for surface expression in vivo. In this study we assess the capacity of FcγRIa–γ-chain complexes expressed in IIA1.6 cells to trigger phagocytosis and ADCC. An intact immunoreceptor tyrosine-based activation motif (ITAM) signalling motif proved essential for triggering of biological function via the FcγRIa receptor complex. Both the FcR γ-chain and the FcγRIIa–ITAM proved active in directing phagocytosis of Staphylococcus aureus and ADCC of erythrocytes, triggered by the FcγRIa complex. The capacity of FcγRIa to trigger phagocytic and cytolytic activity by IIA1.6 cells, both considered ‘professional phagocyte’ functions, motivated us to re-evaluate the cell lineage and developmental stage of IIA1.6 cells. Although originally described as mouse B lymphocytes, the IIA1.6 cells proved positive for non-specific esterase activity and expressed the CD5 antigen. These combined characteristics place the IIA1.6 cells within a unique CD5⁺ B cell/macrophage lineage, optimally suited for cell biological analyses of phagocyte receptors.     

T lymphocyte development in the absence of Fcϵ receptor Iγ subunit: analysis of thymic-dependent and independent αβ and γδ pathways

During fetal development, early thymocyte progenitors transiently express low affinity Fc receptors for IgG (FcγR) of both FcγRII and III isoforms. Only the FcγRIII isoform requires association of an FcγRIII (CD16) α subunit with an FcϵRIγ homodimer for surface expression. To address the role of FcγR in ontogeny, we studied thymic development in FcϵRIγ^−/− mice. We find that day 14.5 CD4⁻CD8⁻ double-negative (DN) fetal thymocytes of FcϵRIγ^−/− mice express mRNA of both FcγRIIb₁ and FcγRIII. Surface expression of FcγRII/III is readily detected on these cells. It appears that FcγRIIb₁, whose surface expression is FcϵRIγ independent, replaces FcγRIII during thymic development in these animals. Moreover, subsequent development into CD4⁺CD8⁺ double-positive and CD4⁺CD8⁻ and CD4⁻CD8⁺ single-positive subsets appears normal even in the absence of FcϵRIγ. However, alterations were noted in adult animals among the DN αβ TCR⁺ thymocytes and peripheral splenic DN T cells as well as CD8αα⁺ intestinal intraepithelial lymphocytes (iIEL). In contrast to conventional T lymphocytes, which do not express either FcγRIII or FcϵRIγ, DN αβ TCR⁺ thymocytes and extrathymically derived αβ TCR⁺ and γδ TCR⁺ CD8αα⁺β⁻ iIEL express TCR which incorporate FcϵRIγ as one of their subunits. Consistent with this, the TCR levels of these cells are lower than the TCR levels on cells from wild-type C57BL/6 mice. Despite the reduction in the level of surface TCR, the development of these cells was unaltered by the absence of FcϵRIγ. Thus, we observed alterations in adult DN αβ TCR⁺ thymocytes, splenic DN αβ TCR⁺ and DN γδ TCR⁺ large granular lymphocytes (LGL), and αβ TCR⁺ and γδ TCR⁺ CD8αα⁺β⁻ iIEL, but no detectable changes in their major fetal thymic developmental pathways. Cultivation of peripheral DN αβ TCR⁺ and DN γδ TCR⁺ cells from FcϵRIγ^−/− mice with interleukin-2 generates LGL which mediate natural killer activity. Unlike LGL from wild-type C57BL/6 mice, LGL from FcϵRIγ^−/− mice lack FcγRIII expression and could not mediate antibody-dependent cellular cytotoxicity through FcγRIII.     

Identification of IgG-binding site on cell membrane

did not bind significantly ertthrocytes (E) but formed a high percentage of rosettes with bovine E sensitized by rabbit IgG (EA_G) or IgM (EA_M) antibody as well as rosettes with human E coated by human anti-CD antibody (EA_CD). Although a weak phagocytosis of untreated E was recorded both types of E were more frequently ingested upon incubation at 37°C when coated with corresponding antibodies. Attachment of EA to amoeba membrane was visualised even after incubation at 4°C or when amoeba cells were fixed with formaldehyde. Binding of sensitized erythrocytes to amoeba surface was markedly reduced when: (i) bovine E were coated with Fab fragment of rabbit IgG antibody; (ii) EA were previously treated with protein A of ; (iii) amoeba were pretreated with monomeric or polymeric IgG or its Fc fragment. It was concluded that possesses Fc-like receptors on its surface.     

New ultrastructural observations: Parallel tubular arrays in human Tγ lymphoid cells

T_γ cells are E-rosetting cells bearing Fc receptors for IgG (E⁺, Fc_γ⁺ cells). Third population (non-T, non-B) lymphoid cells are also Fc_γ⁺ cells and contain unique inclusions called parallel tubular arrays (PTA). Although T_γ cells and third population lymphoid cells should belong to a similar population of cells, previous ultrastructural studies on purified T_γ cells have failed to reveal the presence of PTA. In this study, we have unequivocally demonstrated PTA in the majority of T_γ cells using simple rosetting techniques. A total of 76 EA_hu-rosettes and 108 EA_ox-rosettes prepared from an E⁺ enriched fraction (using sheep erythrocytes as marker particles) were directly examined by electron microscopy. PTA were found in 87% of the EA_hu-rosettes and 82% of the EA_ox-rosettes. Ammonium chloride, commonly used in other laboratories to lyse erythrocytes during the purification procedure was found to cause a marked decrease in the number of ultrastructurally distinct PTA profiles. In contrast, hypotonic lysis had no effect on cellular ultrastructure. This study showed for the first time that T_γ cells are ultrastructurally similar to other Fc_γ⁺ lymphoid cells and contain PTA as a distinct marker. The significance of our findings to the basic function of this E⁺Fc_γ⁺ lymphoid population is discussed.     

The extended hinge region of IgG3 is not required for high phagocytic capacity mediated by Fcγ receptors,but the heavy chains must be disulfide bonded

Feγ receptor (FcγR) phagocytosis and respiratory burst were induced by chimeric mouse-human anti-(4-hydroxy-5-iodo-3-nitrophenyl) acetyl IgG3 antibodies with mutations in hinge and/or in C_H1 region. IgG3 mutants with different hinge length ranging from 47 to 0 amino acids, an IgG3 molecule with an artificial hinge of just one cysteine residue (HM-1), and two hybrid IgG3 molecules with IgG4 hinge or IgG4 C_H1-hinge were tested. Using the monocytic cell line U937 as effector cells, the mutated IgG3 molecules were very similar, revealing high activity, while the IgG3/IgG4 hybrids revealed a slightly reduced activity. However, the hingeless (0-h) mutant was negative, except after interferon-γ stimulation when it became slightly positive. Interestingly, HM-1 was as active as the IgG3 mutants. With polymorphonuclear leucocytes (PMN) as effector cells we obtained some day-to-day variations, but all the IgG3 mutants were highly active, with the two shortest hinge mutants somewhat less active The IgG3/IgG4 hybrid molecules revealed an intermediate activity, while IgG4 wild-type and the 0-h mutant were negative. However, the HM-1 molecule revealed an activity similar to that of the IgG3 mutants. The phagocytic activity of U937 was inhibited by monomeric IgG, indicating the importance of FcγR III. In contrast, with PMN both blockage of FcγRII and cleavage of FcγRIII were required to significantly reduce the phagocytosis and respiratory burst, thus showing that both receptors contribute to the effect. These results demonstrate that the extended IgG3 hinge region is not necessary for a high phagocytic activity and that the major structural importance of the hinge is to connect the two heavy chains in this region.     

Human Fcγ receptors compete for TGN1412 binding that determines the antibody's effector function

The first‐in‐human clinical trial of the CD28‐specific monoclonal antibody (mAb) TGN1412 resulted in a life‐threatening cytokine release syndrome. Although TGN1412 was designed as IgG4, known for weak Fc:Fcγ receptor (FcγR) interactions, these interactions contributed to TGN1412‐induced T‐cell activation. Using cell lines (TFs) expressing human FcγRI, ‐IIa, ‐IIb, or ‐III, we show that TGN1412 and TGN1412 as IgG1 and IgG2 are bound by FcγRs as it can be deduced from literature. However, upon coculture of TGN1412‐decorated T cells with TFs or human primary blood cells, we observed that binding capacities by FcγRs do not correlate with the strength of the mediated effector function. FcγRIIa and FcγRIIb, showing no or very minor binding to TGN1412, mediated strongest T cell proliferation, while high‐affinity FcγRI, exhibiting strong TGN1412 binding, mediated hardly any T‐cell proliferation. These findings are of biological relevance because we show that FcγRI binds TGN1412, thus prevents binding to FcγRIIa or FcγRIIb, and consequently disables T‐cell proliferation. In line with this, FcγRI^?FcγRII⁺ but not FcγRI⁺FcγRII⁺ monocytes mediate TGN1412‐induced T‐cell proliferation. Collectively, by using TGN1412 as example, our results indicate that binding of monomeric IgG subclasses does not predict the FcγR‐mediated effector function, which has major implications for the design of therapeutic mAbs.     

FcγRIIA polymorphism as a risk factor for invasive Streptococcus pneumoniae

Polymorphisms of human Fc gamma receptor IIA (FcγRIIA) have been described and shown to be associated with susceptibility to and severity of certain infectious diseases. Invasive Streptococcus pneumoniae infection continues to be a major cause of morbidity and mortality throughout the world and effective host defense against S. pneumoniae depends on immunoglobulin (Ig) G2-mediated phagocytosis of the bacteria by polymorphonuclear leukocytes. One of the major functions of the FcγRIIA receptor is to play a crucial role in the phagocytosis of IgG2-opsonized bacteria because it is the only receptor able to interact with IgG2 immune complexes. The FcγRIIA polymorphism (FcγRIIA-R131 vs. FcγRIIA-H131) determines the capacity of IgG2-mediated phagocytosis via this receptor. Thus, studies that have examined the direct functional role of R131 and H131 in phagocytosis of the opsonized S. pneumoniae by effector cells in clinically relevant patient groups would provide compelling evidence linking this polymorphism with disease. Here we review the role of FcγRIIA polymorphisms as a host-genetic factor influencing S. pneumoniae infection and describe the in vitro and clinical studies that support the importance of this association.     

Fcγ receptor‐mediated antigen uptake by lung DC contributes to allergic airway hyper‐responsiveness and inflammation

During asthma, lung DC capture and process antigens to initiate and maintain allergic Th2 cell responses to inhaled allergens. The aim of the study was to investigate whether allergen‐specific IgG, generated during sensitization, can potentiate the acute airway inflammation through Fcγ receptor (FcγR)‐mediated antigen uptake and enhance antigen presentation resulting in augmented T‐cell proliferation. We examined the impact of antigen presentation and T‐cell stimulation on allergic airway hyperresponsiveness and inflammation using transgenic and gene‐deficient mice. Both airway inflammation and eosinophilia in bronchoalveolar lavage fluid were markedly reduced in sensitized and challenged FcγR‐deficient mice. Lung DC of WT, but not FcγR‐deficient mice, induced increased antigen‐specific CD4⁺ T‐cell proliferation when pulsed with anti‐OVA IgG immune complexes. Intranasal application of anti‐OVA IgG immune complexes resulted in enhanced airway inflammation, eosinophilia and Th2 cytokine release, mediated through enhanced antigen‐specific T‐cell proliferation in vivo. Finally, antigen‐specific IgG in the serum of sensitized mice led to a significant increase of antigen‐specific CD4⁺ T‐cell proliferation induced by WT, but not FcγR‐deficient, lung DC. We conclude that FcγR‐mediated enhanced antigen presentation and T‐cell stimulation by lung DC has a significant impact on inflammatory responses following allergen challenge in asthma.     

Identification of IgG-binding site on E.-histolytica cell membrane

$\text{E.histolytica}$ did not bind significantly ertthrocytes (E) but formed a high percentage of rosettes with bovine E sensitized by rabbit IgG (EA_G) or IgM (EA_M) antibody as well as rosettes with human E coated by human anti-CD antibody (EA_CD). Although a weak phagocytosis of untreated E was recorded both types of E were more frequently ingested upon incubation at 37°C when coated with corresponding antibodies. Attachment of EA to amoeba membrane was visualised even after incubation at 4°C or when amoeba cells were fixed with formaldehyde. Binding of sensitized erythrocytes to amoeba surface was markedly reduced when: (i) bovine E were coated with Fab fragment of rabbit IgG antibody; (ii) EA were previously treated with protein A of $\text{S.aureus}$; (iii) amoeba were pretreated with monomeric or polymeric IgG or its Fc fragment. It was concluded that $\text{E.histolytica}$ possesses Fc-like receptors on its surface.     

Isotype specificity of Fcγ-receptors on guinea-pig T lymphocytes and their modulation by homologous immune complexes

A previous study has shown that guinea-pig splenic Tγ lymphocytes bearing receptors for guinea-pig Fcγ₁ or Fcγ₂ immunoglobulins delineate two distinct T-cell phenotypes (Ricardo, 1980). In this study, the proportion of splenic Tγ₁ and Tγ₂ lymphocytes were found to vary considerably among different guinea-pigs. Furthermore, the binding of homologous IgG1 soluble immune complexes to guinea-pig splenic Tγ₁ cells led to the modulation of their Fcγ₁-receptors, while the Fcγ₂-receptors on guinea-pig splenic Tγ₂ cells were not affected. The re-appearance of Fcγ₁-receptors after in vitro modulation was detected by inhibiting their EA-rosetting capabilities with homologous aggregated purified IgG1. Following modulation, re-appearance of EA-rosetting by Tγ₁ cells was first detected in 48-hr cultures and by 72 hr their numbers had plateaued to half that present in T-lymphocyte preparations not exposed to immune complexes. Thus, the re-appearance of Tγ₁ cells was slow and some Tγ₁ cells may have irreversibly lost their Fc-receptors subsequent to binding IgG1 immune complexes. Conversely, when Tγ₂ cells were exposed to homologous IgG2-soluble immune complexes only Fcγ₂ receptors were modulated. The pattern of their re-appearance followed that of the modulated Fcγ₁-receptors. These results demonstrated that Fc-receptors on guinea-pig splenic Tγ lymphocytes display isotype specificity for homologous IgG1 and IgG2 molecules and that these receptors can be modulated independently. The presence of isotype-specific Fc-receptors on guinea-pig Tγ lymphocytes could be an important surface determinant involved in the regulation of immune reactions mediated by homologous IgG1 and/or IgG2 antibodies.