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.     

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.     

Characterization of Fc receptors for IgE on human alveolar macrophages.

Human alveolar macrophages (aM phi) isolated from lung lavages performed during bronchoscopy and after surgical removal of pulmonary lobes were analysed for Fc receptors for IgE (Fc epsilon R) and IgG (Fc gamma R) by rosette assays. A mean+/-s.d. of 8.0+/-2.6% of aM phi formed rosettes with fixed ox erythrocytes coated with an IgE myeloma protein (Eo''-IgE). The Eo''-IgE rosettes were inhibited by two IgE myeloma proteins and by IgE Fc fragments but not by myeloma proteins of the other Ig classes or by IgE denatured by heating or reduction and alkylation. Fresh ox erythrocytes sensitized with rabbit IgG antibodies (EoA) formed rosettes with 64.1+/-20.3% of the aM phi. Peripheral blood monocytes formed 10.6+/-1.2% Eo''-IgE and 90.2+/-6.0% EoA rosettes. Incubation of the aM phi with a goat antiserum to human lymphocyte Fc epsilon R inhibited Eo''-IgE rosette formation on aM phi by 80% but did not affect the percentage of EoA rosettes. The antiserum also inhibited Eo''-IgE rosettes formed by peripheral blood monocytes and cultured macrophage-like U937 cells but not those formed by basophilic granulocytes obtained from a patient with chronic myelogenous leukaemia. There was no relationship between age, sex, diagnosis or smoking history of the patients and the percentage of aM phi forming Eo''-IgE rosettes. These studies demonstrate that a subpopulation of human aM phi bear Fc epsilon R that share antigenic determinants with Fc epsilon R on lymphocytes and monocytes. Fc epsilon R(+) aM phi may play an important role in allergic and inflammatory pulmonary diseases by inducing the release of mediators of inflammation after interaction with IgE immune complexes.     

Receptors for IgG (Fc receptors) on human lymphocytes: re-evaluation by multiple techniques

This study examines whether receptors for IgG (Fc receptors), as identified by different methods, are found on identical human lymphocyte subpopulations, and the relationship of Fc receptors to surface immunoglobulin (SIg) and receptor for complement (C'). Fc receptors were identified by two rosetting techniques (antibody-sensitized human erythrocytes, HuEA, or sheep erythrocytes, ShEA) and two immunofluorescent techniques (heat-aggregated IgG of human origin, HuAgg, or of guinea-pig origin, GPAgg).

When lymphocytes depleted of cells rosetting with ShEA were compared to HuEA-depleted lymphocytes, the two subpopulations appeared to be significantly different. However, when lymphocytes were depleted of cells rosetting with ShEA which had been sensitized with lower concentrations of antibody, the subpopulation so depleted appeared to be virtually identical to HuEA-depleted lymphocytes. These studies suggest that more than one lymphocyte subpopulation has Fc receptors and that each subpopulation can, in part, be identified and distinguished from the other by the capacity to bind IgG at differing concentrations.

In particular, these experiments may serve to resolve the controversy concerning the presence of Fc receptors on lymphocytes bearing surface immunoglobulin (SIg). Depletion of lymphocytes rosetting with ShEA removed most of the SIg-bearing lymphocytes; depletion of cells rosetting with ShEA which had been sensitized with lower concentrations of IgG antibody, however, failed to deplete SIg-bearing lymphocytes even though other Fc-bearing populations were completely depleted. These results suggest that SIg-bearing lymphocytes (B lymphocytes) do have Fc receptors but that high concentrations of IgG are needed to demonstrate them.

     

Heterogeneity of human lymphocyte Fc receptors. I. Differential susceptibility to proteolysis

To study the possible heterogeneity of human lymphocyte Fc receptors, isolated human peripheral blood lymphocytes (PBL) were enzymatically altered (`stripped') by exposure to pronase or papain. Pronase treatment markedly increased the percentages of PBL binding IgG-sensitized erythrocytes (EA), while simultaneously removing or inactivating their receptors for heat-aggregated IgG (aggG). Papain treatment markedly diminished the ability of PBL to bind both EA and aggG. Essentially identical results were obtained utilizing EA composed of either human Rh-positive type O erythrocytes sensitized with the human anti-Rh serum Ripley (HRBC-A Ripley) or with chicken erythrocytes sensitized with rabbit anti-CRBC IgG (CRBC-A). CRBC sensitized with Fab'₂ fragments of rabbit anti-CRBC IgG were incapable of forming rosettes with normal or with pronase- or papain-stripped PBL. Pre-treatment of normal lymphocytes with aggG totally ablated their ability to rosette with EA.

Incubation of pronase-stripped PBL for 18–20 hr in 5% CO₂-air at 37°C resulted in diminution (to levels originally present) in the percentages of lymphocytes binding EA, but no regeneration of aggG receptors. Similar incubation of papain-stripped PBL resulted in significant reappearance of receptors binding EA, but no regeneration of aggG receptors. These results strongly suggest that: (1) lymphocyte receptors that bind EA complexes differ from those that bind aggG; (2) some lymphocytes possess cryptic receptors for EA that are expressed after proteolysis with pronase; (3) PBL having receptors for EA also have aggG receptors; and (4) there is no evidence that proteolytic stripping of PBL results in the generation of functionally different receptors for complexed IgG, since the Fc specificity of this receptor remains unchanged.

     

Cytochemistry of Human T-Cell Subpopulations

Acid phosphatase and esterase cytochemistry performed on purified normal human T-cell populations showed that both methods produced distinctive localized dot patterns of reactivity in 60–70% of cells. By examination of rosette preparations formed with ox erythrocytes coated with IgM (EAM), with IgG (EAG), or anti-human κ and λ light chains, it was shown that this pattern of reactivity was largely restricted to small T lymphocytes possessing receptors for the Fc of IgG (Tμ cells). In addition, both B lymphocytes and T cells with receptors for the Fc of IgG (Tλ cells) were larger lymphocytes with more abundant cytoplasm and usually displayed scattered granular acid phosphatase activity; in esterase preparations both cell types were either negative or possessed similar scattered granular positivity. As compared with Tμ cells, Tγ cells were seen to form loose spontaneous rosettes with sheep erythrocytes. Combined esterase and acid phosphatase staining showed that both enzyme activities in the Tμ. cells are localized in the same area, and ultrastructural acid phosphatase cytochemistry established that this was in distinctive lysosomal structures. Tμ staining by both esterase and acid phosphatase cytochemistry was greatly reduced after rosetting with EAG, but not after rosette formation with EAM or sheep erythrocytes.     

Human thymus cells: a study of rosette-forming capacity with unsensitized and sensitized erythrocytes of various species.

Human thymus cells from 117 donors aged 2 months to 15 years, were studied for rosette-forming capacity with unsensitized and sensitized erythrocytes of various species. Viability of the thymus cells at the completion of isolation ranged from 95 to 99%. Over 90% of the total population of thymus cells were identified as T lymphocytes by either standard Es-rosette assay utilizing a 4 degree incubation with preceding centrifugation or T--LCL assay, on the average. Gravity Es-rosette assay high affinity Es-rosette assay, stable Es-rosette assay or active Es-rosette assay for identification of subsets of T lymphocytes indicate that most of the human thymus cells were capable of forming rosettes with unsensitized sheep erythrocytes at higher degrees of temperature with or without preceding centrifugation. Approximately half of the human thymus cells were also capable of forming rosettes with unsensitized allogeneic erythrocytes. Less than 3% of the thymus cells expressed receptors for the Fc portion of IgG (T gamma suppressor cells) whereas less than 1% of thymus cells expressed receptors for the Fc portion of IgM (T mu helper cells), indicating that most of the thymus cells are non-T mu/non T gamma cells. Only 1% or less of the thymus cells were identified as B cells by the SIg assay, the Em-rosette assay or the Ia-like antigen assay. There was no significant difference of any rosette assay between males and females or among various age groups.     

Fc Receptor-Bearing Lymphoid Cells in the Chicken I. Characterization of the Fc(IgG) Receptor

The receptor for Fc(IgG) on chicken lymphoid cells was investigated by EA rosette techniques using sheep erythrocytes sensitized with a subagglutinating dose of anti-sheep erythrocyte (SRBC) chicken serum. Chicken lymphocytes did not form rosettes with SRBC coated with rabbit antibody, and human and mouse lymphocytes did not bind SRBC sensitized with chicken antibody. Only avian sera were effective in blocking the Fc receptor. Similarities between chicken and mammalian Fc receptors were demonstrated as both are pronase sensitive, trypsin resistant, and are distinct from surface immunoglobulin. Fc receptors were also distinguished from avian bursa-and thymus-specific antigens. Additional Fc receptor-bearing cells were revealed in bursa, spleen and bone marrow lymphocytes after neura-minidase treatment.     

Fc Receptor-Bearing Lymphoid Cells in the Chicken I. Characterization of the Fc(IgG) Receptor

The receptor for Fc(IgG) on chicken lymphoid cells was investigated by EA rosette techniques using sheep erythrocytes sensitized with a subagglutinating dose of anti-sheep erythrocyte (SRBC) chicken serum. Chicken lymphocytes did not form rosettes with SRBC coated with rabbit antibody, and human and mouse lymphocytes did not bind SRBC sensitized with chicken antibody. Only avian sera were effective in blocking the Fc receptor. Similarities between chicken and mammalian Fc receptors were demonstrated as both are pronase sensitive, trypsin resistant, and are distinct from surface immunoglobulin. Fc receptors were also distinguished from avian bursa-and thymus-specific antigens. Additional Fc receptor-bearing cells were revealed in bursa, spleen and bone marrow lymphocytes after neura-minidase treatment.     

Surface markers of porcine lymphocytes and distribution in various lymphoid organs.

Three rosette test systems were developed for the classification of subpopulations of porcine lymphocytes and correlated with the presence or the absence of membrane immunoglobulins, as well as agg-P-IgG receptors, by means of various RFC depletion experiments. Among the ED-RFC negative cells, PBL with surface immunoglobulins (SIg), at 4 degrees C, can be subdivided into two about equal populations: (1) One half of these cells have surface stable SIg (B cells) and mu determinants; they possess C3b receptors leading to the formation of rosettes with complement-coated zymosan particles (ZC). (2) The other half have surface labile Ig when the cells are washed at 37 degrees C (L-cells); they possess Fc receptors as detected by EA-RFC, i. e. pig-IgG-coated ox RBC. Receptors for Fc-agg, as detected by agg-P-IgG, were primarily shown on B cells. ZC and EA-RFC are in the highest tissue concentration, in bone marrow and spleen, respectively, and more numerous in adults than in newborns. Moreover, the number of EA-RFC was twofold higher than that of ZC-RFC in spleen, mesenteric lymph node and cisterna chyli. This latter organ was the only one without null cell.     

Characterization of the Fc(IgG) receptor on the pluripotential leukaemia cell K-562.

K-562 cells express an Fc receptor that is murine isotype IgG specific. The receptor was defined by rosette formation using sheep erythrocytes sensitized with murine monoclonal haemagglutinin (EA) of known isotype. Optimal rosette formation occurred at 4 degrees C or ambient temperature; however, the number of rosettes formed at ambient temperature decreased after 8 h whereas formed rosettes were stable at 4 degrees C. EA in which IgG2b was the immunoglobulin isotype gave high numbers of rosettes while IgG2a gave lower but significant numbers. Aggregated human IgG inhibited rosette formation of EA(IgG2a) more easily than those of EA(IgG2b), indicating a higher affinity of the Fc receptor for IgG2b.     

Studies on the nature of EA binding by lymphocytes from rheumatoid arthritis patients

Investigation of the nature of the increased erythrocyte-antibody (EA) binding activity of peripheral blood lymphocytes (PBL) from rheumatoid arthritis (RA) patients reported in the preceding paper has revealed that IgG is the active class of antibody in this rosette formation. Some IgM binding also occurs. SRBC sensitized with F(ab)₂ preparations of IgG do not give rosette formation even at high concentrations. EA binding is inhibited by prior incubation of lymphocytes with heat-aggregated human IgG but antigen-antibody complexes did not give significant inhibition.The majority of these rosettes were found to be stable at 4°C and room temperature but labile at 37°C.Enzyme studies with pronase, trypsin, neuraminidase and treatment with sodium azide gave results strongly supporting the conclusion that the increased binding observed is increased Fc-receptor activity. This activity appears not to be a result of Fc binding by cell-bound rheumatoid factor.A range of titres of antibody and of IgG was used to sensitize erythrocytes to form EA and the enhanced EA-rosette formation by PBL from RA patients occurred throughout the range of concentrations of sensitizing antibody. Significantly more EA were bound by individual lymphocytes from RA patients than control subjects. This data suggest that the Fc receptors on RA lymphocytes are more avid for EA than receptors on lymphocytes from healthy people.     

The functional activity of Fc gamma RII and Fc gamma RIII on subsets of human lymphocytes.

Subsets of human lymphocytes were isolated from peripheral blood using magnetic beads coated with anti-CD4, -CD8, -CD19 or -CD56 antibodies to yield T4, T8, B and natural killer (NK) cell suspensions with greater than 95% purity. The functional activity of Fc gamma receptor II (Fc gamma RII) and Fc gamma receptor III (Fc gamma RIII) on these subsets was assessed by measuring rosette formation with red cells sensitized with known levels of either rabbit IgG or human (monoclonal or polyclonal) IgG1 anti-D, IgG3 anti-D or IgG3 anti-c (E-IgG). Lysis of red cells by K cells (mediated by Fc gamma RIII) in antibody-dependent cell-mediated cytotoxicity (ADCC) assays was promoted by polyclonal and some monoclonal antibodies. Using these 'ADCC+' antibodies, minimum red cell sensitization levels required to promote rosette formation with NK cells were 2000 IgG1 or IgG3 molecules/red cell compared to 15,000 IgG1 or 4000 IgG3 molecules/red cell with 'ADCC-' monoclonal antibodies. The greater efficiency of ADCC+ antibodies is consistent with their previously reported ability to bind Fc gamma RIII via CH2 and CH3 domains whereas ADCC- antibodies bind only via CH3 domains. B cells formed rosettes only at high levels of sensitization: approximately 60,000 IgG1 or 20,000 IgG3 anti-D molecules/cell. These data reflect the low affinity of Fc gamma RII for monomeric human IgG. Although over 90% of NK cells bound anti-CD16, and 70% formed rosettes with red cells sensitized with rabbit IgG (30,000 molecules/cell), only 25% of NK cells formed rosettes with E-IgG3 at 100,000 IgG molecules/cell. Approximately 35% of B cells, 10% of T8 cells but no T4 cells formed rosettes with E-IgG (100,000 IgG3 molecules/cell). With T8, B and NK cells, IgG3 anti-D promoted greater rosette formation than IgG1 anti-D at comparable levels of sensitization. Presumably the longer hinge region of IgG3 enabled it to bridge the gap between negatively charged lymphocytes and red cells more efficiently than IgG1.     

The mechanism of rosette formation between Rh (D) positive erythrocytes and peripheral blood lymphocytes from Rh isoimmunized individuals. The role of surface micro-projections

Peripheral blood leucocytes were obtained from Group A Rh (D)-negative male volunteers re-injected with A Rh (D)-positive erythrocytes and the mechanism by which they form rosettes in vitro with A Rh (D)-positive erythrocytes was studied. It was found that there was a marked reduction in the number of rosettes formed at 4° as compared with 37°. Moreover, those rosettes formed at 4° comprised a central rosette-forming cell surrounded by one layer of erythrocytes whereas at 37° some rosette-forming cells (RFC) appeared to be surrounded by agglutinated erythrocytes. No anti-D antibody-forming cells were detected by the localized haemolysis in gel technique despite the presence of large numbers of RFC in the preparations tested. Pretreatment of leucocyte aliquots with anti-sera specific for human Fab at 4° was nearly as effective as pretreatment with anti-Fab at 37° in inhibiting rosette formation.

Examination of serial sections of rosettes with the electron microscope revealed that erythrocytes were often attached to micro-projections from the surface of central RFC. Cytochalasin B partially inhibited rosette formation at 37°. Those rosettes formed in the presence of cytochalasin B consisted of a central rosette-forming cell with few adherent erythrocytes and with the electron microscope it was seen that erythrocytes were rarely attached to central RFC by micro-projections. It is suggested that in this system the contractile activity of the surface of rosette-forming cells is important in rosette formation.

     

Shedding of Fc receptor from mononuclear cells and their ability of binding IgG1 and IgG3 anti-Rh/D antibodies

Fc receptors for IgG1 and IgG3 on peripheral blood lymphocytes and monocytes were studied before and after temperature shift from 4-37 degrees C. The investigations were performed in the EA test using human erythrocytes sensitized with anti-Rh/D/antibodies of IgG1 (EA IgG1) and IgG3 (EA IgG3) subclasses. It occurred that lymphocytes and monocytes were able to bind IgG1 and IgG3 antibodies before and after shedding, however, lower percentage of rosette was observed after temperature shift. This decrease was similar in the EAIgG1 and EAIgG3 tests. The supernatants obtained during shedding occurred to contain active Fc receptors since the inhibition of rosette formation was obtained after the incubation of sensitized erythrocytes with these supernatants. IgG1 as well as IgG3 myeloma proteins inhibited rosette formation in both EAIgG1 and EAIgG3 tests. Our data might suggest that IgG1 and IgG3 anti-D antibodies are able to bind to the same Fc receptor on lymphocytes as well as on monocytes.     

Active and passive re-expression of Fcγ receptors on human lymphocytes

Modulation of receptors for IgG (FcγR) on human lymphocytes was induced by the interaction with erythrocyte-IgG antibody (EA_G) complexes followed by incubation at 37°C. The re-expression of FcγR could be achieved by two independent processes, (a) Active synthesis, susceptible to inhibition by puromycin or cycloheximide was shown to peak 4 to 6 h after removal of EA_G complexes; it required addition of at least 2% fetal calf serum. (b) Insertion of soluble FcγR into the membrane of modulated lymphocytes was shown to occur within 20 min of contact between cells and FcγR-containing supernatants; it was not altered by protein synthesis inhibitors. FcγR-like material, spontaneously released by unstimulated peripheral blood lymphocytes or by polymorphonuclear cells, was taken up by modulated lymphocytes. This soluble material was fully absorbed on polymerized human IgG; it was non- dialyzable, thermolabile (56 °C, 30 min) and partially destroyed by freezing and thawing; it was recovered as two broad peaks from chromatography on polyacrylamide gel; it was shown to bind to both T and non-T FcγR-bearing lymphocytes capable of forming EA_G rosettes before modulation; and it could be inserted into allogeneic lymphocytes. These results demonstrate that the FcγR structure bears two active sites, one binding to the Fcγ and the other to the surface of EA_G rosette-forming cells. Rapid release of soluble FcγR from the cells and their possible insertion into the membrane may have important implications with respect to the biological functions associated with these receptors.     

The Fc Receptors of Normal and Cancer Patients Monocytes

The ability of human monocytes from normal donors and gastric-cancer patients to form rosettes with ?0? Rh⁺(D) human erythrocytes coated with hyperimmune IgG anti-D antibody (EA_hu) and to kill the same target in antibody-dependent cellular cytotoxicity (ADCC) were assessed. Trypsin pretreatment of normal monocytes decreased their ability to form rosettes with EA_hu complexes, but their ADCC activity was unaffected. The Fc receptor (FcR) expression and ADCC activity of monocytes of cancer patients were elevated, and trypsin-treatment led to their further increase. The elevated values were related to the presence of the tumour. These results may suggest that human monocytes possess trypsin-sensitive and trypsin-resistant Fc receptors. The trypsin-resistant FcR seems to be involved in ADCC phenomenon and to be preferentially expressed on monocytes of some cancer patients.     

Lymphocyte subpopulations in peripheral blood of healthy persons. Characterization by surface markers and lack of selection during purification

Lymphocytes were isolated at 99% purity from peripheral blood of healthy persons by defibrination, gelatine sedimentation, treatment with carbonyl iron powder and centrifugation on Ficoll–Isopaque. Subpopulations were identified by three surface markers: cells forming rosettes with sheep red blood cells (SRBC) (E-binding lymphocytes) as a measure of T lymphocytes; lymphocytes with surface immunoglobulin identified by indirect immunofluorescence (B lymphocytes); lymphocytes with receptors for C3 observed by the rosette method using SRBC treated with rabbit antiserum and human complement (EAC-binding lymphocytes).

The yield of lymphocytes after purification varied from 15 to 65%. No selection of lymphocytes was observed either by counting immunoglobulin-bearing and EAC-binding lymphocytes in whole blood and in purified cells from the same sample, or by statistical analysis of lymphocytes in subpopulations as a function of the yields from twenty-six experiments. In the absence of selection during purification the total numbers of T and B lymphocytes could be calculated from the percentages and the total numbers of lymphocytes. Our normal values are close to those reported using other non-selective methods of purification.

When lymphocytes were simultaneously stained for immunoglobulin and rosetted with EAC, cells bearing either or both markers were found. In total, 27–35% cells were identified by these markers. Since about 70% of the cells were E-binding, practically all lymphocytes could be identified. A small overlap between E-binding and immunoglobulin-bearing/EAC-binding lymphocytes may occur.

Either the IgM or the IgG-containing fractions obtained after fractionation of rabbit anti-SRBC serum on Sephadex G-200 could be used for sensitization of SRBC with complement. Formation of rosettes was not prevented by pretreating the lymphocytes with aggregated IgG, while rosettes formed with EA prepared by high concentrations of IgG antibody (Fc-binding lymphocytes) were abolished. It is concluded that rosettes formed with IgG-EAC (or whole serum EAC) using diluted antiserum identify complement-reactive lymphocytes and are not caused by synergism with Fc receptors. When SRBC were sensitized with varying dilutions of whole antiserum or its IgG fraction identical plateaus for the percentages of EAC-binding lymphocytes were found. Subagglutinating concentrations of the IgM fraction was insufficient to reach the plateau and also consistently resulted in lower values for EAC-binding lymphocytes.

     

Which Fc receptor-bearing cells are detected with the Ripley assay?

Mononuclear cells were isolated from peripheral blood and analysed with T and B markers (E rosettes and SIg) and on contaminating monocytes and PMN. The suspensions contained 63.3 +/- 4.8% T lymphocytes, 11.4 +/- 3.8% B lymphocytes, 9.0 +/- 5.5% null lymphocytes, 15.1 +/- 3.8% monocytes and 1.1 +/- 0.6% PMN. Of all cells, 27.6 +/- 12.1% formed EA rosettes with OR1R2 red cells coated with anti-CD Ripley. In preparations fixed after cytocentrifugation, the EA rosette-forming cells were studied with regard to esterase activity. The proportion of cells with detectable Fc receptors was further studied in purified T lymphocyte and in monocyte suspensions. Finally, EA rosettes were isolated by gradient centrifugation and the rosette forming cells studied with conventional T and B markers. All procedures gave corresponding results: on average 11-14% of the T lymphocytes and nearly 100% of the null cells formed EA rosettes, while only 2% of the B lymphocytes had detectable Fc receptors. Of the purified monocyte and PMN populations, on average 72 +/- 10.5 and 14.5 +/- 5.4%, respectively, formed EA rosettes. Thus, the Ripley assay, representing an important marker for null lymphocytes, cannot be regarded as specific for this population of white blood cells.     

T and B lymphocytes in the marmoset: a natural haemopoietic chimera.

The thymus-derived (T) lymphocyte and bone marrow-derived (B) lymphocyte populations of the marmoset were characterized using specific cell surface markers Approximately 85% of the thymocyates formed rosettes with neuraminidase-treated sheep erythrocytes (En). The percentage (approximately 69%) of peripheral blood lymphocytes (PBL) forming rosettes with En was the same as that which stained with fluorescently labelled goat anti-marmoset thymocyte serum (ATS). These two assays identified the same cell population since treatment of cells with ATS and complement resulted in a concomitant decrease in En rosette formation. Marmoset PBL also formed rosettes with human erythrocytes sensitized with antibody and complement (HEAC); since the percentage (approximately 20%) HEAC rosette was the same as that of cells stained with fluorescently labelled goat anti-marmoset IgG, these cells were considered to be B cells. A small percentage of cells (approximately 1-5%) possessed both types of receptors. The mean percentages of T and B cells present in PBL of single-born, presumably non-chimeric animals, were the same as that of iso-sexual and heterosexual chimeras.