Lymphocyte Cell Surface Antigens Research Articles

Lymphocyte subset distribution in apparently normal and single intradermal test-positive water buffaloes analyzed by flow cytometry

Monoclonal antibodies (mAbs) against bovine lymphocyte cell surface antigens namely, MHC Class I, MHC class II (DP, DQ and DR), CD3, CD4, CD8, gamma delta TCR, WC1N1 and WC1N2, were tested for their reactivity on apparently normal buffalo mononuclear cells prepared from spleen, lymph nodes and peripheral blood. All the mAbs cross-reacted with the buffalo mononuclear cells. The mean (±SD) CD4:CD8 cell ratio in the peripheral blood of apparently normal buffaloes was 1.08±0.049 while in the spleen and lymph nodes it was 0.90±0.080 and 1.81±0.430, respectively. The lymphocyte subsets in the buffaloes positive for tuberculosis by the single intra dermal (SID) test was found to be altered; the CD4 cells were reduced while the CD8 and gamma delta cells were increased. The mean CD4:CD8 ratio in the SID positive buffaloes was 0.36±0.010.

Novel assays of multiple lymphocyte functions in whole blood measure: New mechanisms of action of mycophenolate mofetil in vivo

The antiproliferative effects of MMF are believed to be the mechanism of its immunosuppressive action. We further investigated the mechanisms of action by assessing the pharmacodynamics (PD) of MMF in treated animals using whole blood assays not only of lymphocyte proliferation but also of activation. In vitro, different MPA concentrations were added to rat whole blood. In vivo, Lewis rats were treated with single doses of 5, 10 or 20 mg/kg MMF ( n=6 rats/dose group). Blood was obtained before and at different times after drug administration. For both in vitro and in vivo studies, different mitogens with calcium-dependent (TCR) or -independent (co-stimulatory) pathways of lymphocyte activation were added to the blood for stimulation. Proliferation was measured by [ 3H]TdR incorporation and by flow cytometric detection of DNA content. Activation was measured by changes in T cell surface expression of CD25, CD134, CD71, CD11a and CD54. In vitro and in vivo studies showed a dose-dependent inhibition by MPA and MMF, respectively, of lymphocyte proliferation and surface antigen expression. We observed high correlations between MMF PD effects over time with both MMF dose and MPA plasma concentrations in vivo. We show that MMF, apart from its antiproliferative effect, induced a dose-dependent suppression of calcium-dependent and -independent stimulated expression of important lymphocyte cell surface antigens. These data suggest that the ex vivo assessment of immune function in whole blood can uncover new mechanisms of MMF action. Our results demonstrated that the measurement of the PD is a means to assess the functional effects of MMF after its administration in vivo.

CD48-deficient mice have a pronounced defect in CD4(+) T cell activation.

We have generated mice deficient in the expression of the lymphocyte cell surface antigen CD48 (Blast-1, BCM1, sgp-60) by gene targeting in embryonic stem cells. Mice homozygous for the CD48 mutation (CD48(-/-) mice) are severely impaired in CD4(+) T cell activation. Proliferative responses to mitogens, anti-CD3 mAb, and alloantigen are all reduced. Experiments in which T cells and antigen-presenting cells from either wild-type or CD48(-/-) mice were cocultured reveal that CD48 is important on both T cells and antigen-presenting cells. The most dramatic impairment was observed in experiments in which highly purified T cells were stimulated through the T cell receptor in the presence of the phorbol ester, phorbol 12-myristate 13-acetate. The results of these experiments raise the possibility that CD48 plays a role in signaling through the T cell receptor.

Retinoid-Mediated Signaling Pathways in CD38 Antigen Expression in Myeloid Leukemia Cells

The lymphocyte cell surface antigen CD38, which was originally described as a differentiation marker, has emerged as an important multifunctional protein. Its most intriguing and well characterized function is its ability to catalyze the synthesis of cyclic ADP-ribose (cADPR) from NAD. cADPR serves as an important second messenger in controlling the release of intracellular calcium from ryanodine-sensitive intracellular pools. By virtue of its ability to synthesize cADPR as well as to act as an adhesion and signal transduction molecule, CD38 may play a role in such diverse physiological processes as cell growth, apoptosis, differentiation, and inflammation. Equally interesting is the pattern of CD38 expression in hematopoeitic cells. In the bone marrow, early precursor cells predominantly express CD38 antigen, whereas mature circulating blood cells lack or express very low levels. The expression is also high on malignant hematopoeitic cells and thus may be of prognostic relevance in certain leu-kemias. Presently, there is little information available on the factors that regulate the expression of CD38 antigen in hematopoeitic cells. In this review, we summarize recent findings on the regulation of CD38 antigen by retinoids (vitamin A and related compounds). At least in the myeloid cell lineage, retinoids appear to be exquisitely potent and specific inducers of CD38 antigen expression, and retinoid-induced expression of CD38 is mediated via activation of the retinoic acid-alpha (RARα) nuclear receptor.

The major histocompatibility complex of ruminants.

Studies of the major histocompatibility complex (MHC) of cattle over the past twenty years have revealed a reasonably detailed picture of the genetic organisation and function of the genes within this genetic system. Serological and biochemical analysis of lymphocyte cell surface antigens provided the first evidence for highly polymorphic MHC genes in cattle and other ruminant species. The MHC of cattle was thus named the bovine leucocyte antigen (BoLA) system. During the past 10 years, tools of molecular biology have been used to characterise the number of MHC genes, their sequence and fine structure in a number of ruminant species. Although individual MHC genes were found to have clear orthologues among ruminants and other mammalian species, the MHC of cattle, and probably that of sheep and goats, has a unique genetic organisation. Cattle have a class II gene cluster (class IIb region) which is physically distant from all the other MHC genes on the same chromosome. Moreover, genes involved in antigen processing, such as the proteosome subunit locus LMP2, are also found in the class IIb region, demonstrating that these genes need not be in close proximity to other MHC genes to function normally. The MHC class I and class II gene products of ruminants present processed peptides to T lymphocytes which mediate helper and cytotoxic functions. Identification of peptide binding motifs of cattle MHC class I molecules indicates that ruminant MHC molecules function in a similar manner to those of mice and humans. These functional studies provide a firm molecular basis for a number of well-documented associations with infectious diseases, although a detailed understanding of the immunogenetic mechanisms underlying these associations has yet to be elucidated.

Effect of pentachlorophenol on immune function

The organochlorine compound, pentachlorophenol, was evaluated for effects on immune system function in male Fisher 344 rats. Pentachlorophenol was prepared in an olive oil vehicle and was administered by oral gavage twice weekly for 28 days at a dose of 2.0 mg/kg per treatment. Exposure to pentachlorophenol increased body weight gains (P=0.024) during the treatment period. Liver (P=0.034) and kidney (P=0.012) body weight ratios were also increased. Pentachlorophenol exposure enhanced T-lymphocyte blastogenesis induced by concanavalin A (Con A)(P=0.0001) and phytohemagglutinin (PHA)(P=0.048) evaluated using stimulation indices. Corresponding B-lymphocyte blastogenesis induced by lipoploysaccharide/dextran (LPS/dex)(P=0.0034) was also enhanced by pentachlorophenol exposure. Pentachlorophenol suppressed the antibody response against sheep red blood cells (SRBCs) by 39% when the response was expressed per viable spleen cell (P=0.006). This suppression was not evident when the response was expressed per spleen (P=0.22), suggesting that a compensatory mechanism or extramedullary splenic hemopoiesis was occurring minimizing the overall impact on humoral immunity. The enhanced B- and T-lymphocyte blastogenesis may also reflect compensatory or hemopoietic activity. Pentachlorophenol exposure had no effect on peritoneal macrophage phagocytosis (P=0.31) or lymphocyte cell surface antigen expression. The observed alterations in lymphocyte blastogenesis and humoral immunity subsequent to pentachlorophenol exposure do not appear to be associated with phagocytosis or lymphocyte cell surface antigen expression.

Analysis of splenic and thymic lymphocyte subpopulations in chickens infected with Salmonella enteritidis

Lymphocytes expressing CD3, CD4, CD8, pan lymphocyte, IgA, IgG and IgM cell surface antigens were assessed by in the spleen and thymus of chickens following infection with Salmonella enteritidis using flow cytometric analysis. At 6 days post primary infection and 2 days post secondary infection with S. enteritidis, the percentages of IgA + and IgM + lymphocytes in the spleen were significantly increased ( P < 0.05). At 2 days post secondary infection with S. enteritidis, the percentage of CD4 + T lymphocyte in the spleen and CD8 + T lymphocyte percentage in the thymus were significantly increased ( P < 0.05). These results indicate that S. enteritidis infection induces changes in the spleen and thymus that reflect the dynamics of the host protective immune response.

Induction of interleukin-1 and glucocorticoid hormones by HIV promotes viral replication and links human chromosome 2 to AIDS pathogenesis: genetic mechanisms and therapeutic implications

Human immunodeficiency virus may regulate its replication by stimulating the synthesis of interleukin-1. Interleukin-1, in turn, has the ability to stimulate the human immunodeficiency virus enhancer region. The human genes responsible for interleukin-1 and interleukin-1 receptor antagonist synthesis are located on the long arm of chromosome 2. Coincidentally, the trans-activation responsive ribonucleic acid element in the R region of the long terminal repeat of human immunodeficiency virus-1 has been found to interact directly with a factor present on the long arm of chromosome 2 to facilitate transactivation by the human immunodeficiency virus Tat protein. The human CD26 gene is also located on the long arm of chromosome 2. CD26 is a lymphocyte cell surface antigen that is stimulated by interleukin-1 and serves with CD4 as a coreceptor that interacts with the V3 loop in gp120 of human immunodeficiency virus. The human immunodeficiency virus-induced interleukin-1 excess, thus, serves human immunodeficiency virus by enhancing replication, and by increasing human immunodeficiency virus infectivity via activation of CD26. IL-1 also adversely affects acquired immune deficiency syndrome-related Kaposi's sarcoma. Several genetic treatments for human immunodeficiency virus infection are proposed.

In vitro detection of Mycoplasma fermentans binding to B-lymphocytes in fresh peripheral blood using flow cytometry.

Previous reports have shown, using fluorescent probes conjugated to the organism, that Mycoplasma fermentans fuses with about 12% of peripheral blood lymphocytes. However, no lymphocyte subset was specified. To elucidate the specific subset of lymphocytes involved, we developed a three-color flow cytometric assay to detect M. fermentans binding to fresh peripheral blood cells. In our assay, two strains of M. fermentans were grown in SP4 glucose broth, mixed with fresh whole blood samples (n > 20), and incubated at 37 degrees C. The blood samples were then stained with a polyclonal antibody to M. fermentans, a monoclonal antibody to B-lymphocytes (CD19), and a monoclonal antibody to T-lymphocytes (CD3). Using three-color flow cytometry, we obtained data confirming binding of M. fermentans to 10%-15% of peripheral blood lymphocytes with minimal granulocyte or monocyte staining detected. Flow cytometric analysis showed that early binding appears predominantly directed towards B-lymphocytes (86.7 +/- 9.0%), and that this binding could not be blocked by antibodies directed towards common B lymphocyte cell surface antigens. M. fermentans binding to B-lymphocytes occurred within 5 min of in vitro inoculation, reached a maximum within 30-60 min (94-97%), and thereafter plateaued. The binding was concentration dependent over a three log dilution using 10(3) color changing units as standard. Binding to T-lymphocytes was minimal (<5% positive). B lineage tumor cells or peripheral blood B cells obtained from HIV infected individuals demonstrated reduced binding of M. fermentans. This assay provides a good method to study the cellular interactions of mycoplasma and may help to elucidate pathogenic mechanisms of mycoplasma infections.

Immunosuppression with monoclonal antibodies and CTLA4-Ig after myoblast transplantation in mice.

Various combinations of monoclonal antibodies specific for lymphocyte cell surface antigens and a recombinant molecule (CTLA4-Ig) were used to immunosuppress mice after transplantation of MHC-incompatible myoblasts. To assess the effectiveness of the immunosuppression, the donor myoblasts were obtained from a transgenic mouse (TnI LacZ1/29) containing a beta-galactosidase (beta-gal) reporter gene under the control of a muscle-specific promoter. No muscle fibers expressing beta-gal were observed 1 month after the myoblast transplantation, when the animals were not immunosuppressed or were treated with CTLA4-Ig alone. Approximately 50% of the muscle fibers expressed the beta-gal reporter gene 1 month after transplantation in mice treated with CTLA4-Ig combined with an anti-CD4 monoclonal antibody and in mice treated with a combination of anti-CD4, anti-CD8, and anti-lymphocyte function-associated antigen-1. The percentage of beta-gal-labeled muscle fibers increased to 94% when this combination of the three monoclonal antibodies was administrated weekly for 3 weeks. Although excellent graft survival rates were obtained 1 month after transplantation, reflecting an effective immunosuppression by these three treatments, no beta-gal-positive fibers were found 2 months after the transplantation, indicating the inability of these immunosuppressive agents to maintain long-term graft survival and induce tolerance to the myoblasts and muscle fibers of donor origin.

Stimulation of ADP-ribosyl cyclase activity of the cell surface antigen CD38 by zinc ions resulting from inhibition of its NAD+ glycohydrolase activity.

The lymphocyte cell surface antigen, CD38, which has an amino acid sequence similar to Aplysia ADP-ribosyl cyclase, catalyzes not only the hydrolysis of NAD+ and 1-(5-phospho-beta-D-ribosyl) adenosine 5'-phosphate cyclic anhydride (cyclic ADP-ribose) but also the formation of cyclic ADP-ribose from NAD+. To characterize the bifunctional enzyme properties, we produced the recombinant CD38 fused with a maltose-binding protein (MBP-CD38). Zinc ions stimulated the ADP-ribosyl cyclase activity of MBP-CD38, but inversely inhibited its NAD+ glycohydrolase activity which was approximately 100-fold dominant to the cyclase activity in the absence of Zn2+. Such dual effects of Zn2+ were also observed in the native membrane-bound CD38 of HL-60 cells which had been caused to differentiate by retinoic acid. Zinc ions inhibited the NAD+ glycohydrolase reaction catalyzed by MBP-CD38 in an uncompetitive manner, whereas they enhanced the ADP-ribosyl cyclase reaction without affecting the Km value for NAD+. There was an increase in the fluorescence intensity of a hydrophobic fluorescent probe, 8-anilino-1-naphthalenesulfonate, in the presence of MBP-CD38. The fluorescence increase was further enhanced by the addition of Zn2+ with a shift in the maximum emission wavelength from 484 nm to 470 nm, suggesting that Zn2+ caused conformational changes of MBP-CD38. These results indicate that Zn2+ directly interacts with CD38 to stimulate its ADP-ribosyl cyclase with inhibition of its NAD+ glycohydrolase, probably due to prevention of the access of water molecule to an intermediate of the enzymesubstrate complex.

Genomic organization, exact localization, and tissue expression of the human CD26 (dipeptidyl peptidase IV) gene.

CD26 is a lymphocyte cell surface antigen which is increased during T-cell activation and is also expressed in other tissues. It is an atypical serine protease belonging to the prolyl oligopeptidase family. CD26 has been implicated in a variety of biological functions including T-cell activation, cell-to-cell adhesion, and recently in HIV infection. This paper describes, through the isolation and partial sequencing of eight human CD26 genomic clones, the first information on the genomic organization of the prolyl oligopeptidase family. We have established that the human CD26 gene spans approximately 70 kilobases (kb) and contains 26 exons, ranging in size from 45 base pairs (bp) to 1.4 kb. The nucleotides that encode the serine recognition site (G-W-S-Y-G) are split between two exons. This clearly distinguishes the genomic organization of the prolyl oligopeptidase family from that of the classical serine protease family. The 5' flanking domain of the CD26 gene contains neither a TATA box nor a CAAT box, but a 300 bp region extremely rich in C and G (72%) contains potential binding sites for several transcriptional factors. The human CD26 gene encodes two messages sized at about 4.2 and 2.8 kb. These are both expressed at high levels in the placenta and kidney and at moderate levels in the lung and liver. Only the 4.2 kb mRNA was expressed at low levels in skeletal muscle, heart, brain, and pancreas. Fluorescence in situ hybridization on metaphase chromosome spreads located the human CD26 gene to the long arm of chromosome 2(2q24.3).

Interferon Gamma Administration Increases Monocyte HLA-DR Antigen Expression but Not Endogenous Interferon Production

To determine the effect of the adjuvant administration of interferon gamma on monocyte HLA-DR antigen expression and mitogen-stimulated interferon gamma production following injury. Double-blind, randomized, placebo-controlled trial. University Hospital, Newark, NJ, a level I trauma center. Persons older than 16 years with an Injury Severity Score greater than 20 and documented bacterial contamination at the time of injury (N = 98). Recombinant human interferon gamma (n = 46; 0.1 mg subcutaneously) or placebo (n = 52) was given for 10 days following injury. Incidence of major infection, monocyte and lymphocyte cell surface antigen expression, and interferon gamma production at multiple time points following injury. Peripheral monocyte HLA-DR was measured as percent of cells staining positive and as mean channel fluorescence. Both values were significantly increased in the interferon gamma group compared with the placebo group on days 3, 5, 8, and 11. The incidence of major infection was unaffected by interferon gamma administration. Infection decreased percent of HLA-DR-positive monocytes and mean channel fluorescence as compared with noninfected patients on postinjury days 8 and 11 in the placebo group but not in the interferon gamma group. Interferon gamma production improved from 3 +/- 3 U/mL on day 1 to 15 +/- 10 U/mL by day 30 but was always significantly lower than normal (25 +/- 3 [mean +/- SD] U/mL). Interferon gamma production was unaffected by either infection or interferon gamma administration. Interferon gamma administration after injury stimulated monocyte HLA-DR antigen expression and density but failed to improve interferon gamma production, a T-cell-mediated function. The incidence of infection was not decreased by the administration of interferon gamma for 10 days. Improvement in monocyte HLA-DR antigen expression did not correlate with a global restoration of immune function, and other interventions will be necessary to decrease infection after injury.

Characterization of an Early Activation‐Dependent Antigen on Lymphocytes Defined by the Monoclonal Antibody BL‐Ac(F2)

A novel activation-dependent lymphocyte cell-surface antigen which is recognized by a MoAb, BL-Ac(F2), is described. Although not found on resting lymphocytes the antigen is induced rapidly within 2-4 h following stimulation of the cells using mitogens or antibodies against the T-cell CD3 antigen and sIgM on B cells, respectively. Immunoprecipitation and Western Blotting indicated that the MoAb recognizes a molecule in a range of 78-85 kDa. Beyond its activation-dependent expression on lymphocytes the antigen was detected also on myelo-monocytic cells. Expression kinetics and cellular distribution of this molecule suggest that it is distinct from previously described activation-dependent cell-surface antigens such as CD69, CD25 and 4F2.

Immunogold-Silver Staining Method for Lymphocyte Cell Surface Antigens with Light, Transmission Electron, and Scanning Electron Microscopy

Abstract The immunogold-silver staining (IGSS) method combined with light, transmission electron, and scanning electron microscopy (LM, TEM and SEM, respectively) was used for detecting lymphocyte surface antigens. Two different sizes of colloidal gold particles (5 nrn and 15 nm) were applied as markers and IntenSEII kit as a physical developer for gold particles.The silver enhanced gold particles were clearly observed on cell surfaces as black dots in LM and TEM and as white dots in SEM equipped with a mixed signal of secondary electron and back-scattered electron (SE/BE) signals. Monoclonal antibody (MAb)-positive cells possessing the complexes on their well preserved surfaces were easily identified among other lymphoid cells at low magnifications of LM or SEM equipped with SE/BE signals. Thus, the IGSS method has a great advantage for a qualitative screening such as the percentage of lymphocyte subsets in a cell suspension. However, the IGSS method was inadequate for semiquantitative study with antigen...

Generation of monoclonal antibodies to human lymphocyte cell surface antigens using insect cell expressing recombinant proteins

We have expressed human CD40 and human B7 in insect cells using the baculovirus expression system and have used these insect cells to immunize mice for the generation of monoclonal antibodies. We demonstrate here that specific monoclonal antibodies to human CD40 and human B7 were obtained using this approach. One significant advantage of this method is that immunizing mice with insect cells did not evoke an immune response to human cells and, therefore, EBV-transformed human B cells could be used to screen for specific antibody production by the hybridoma clones.

DEVELOPMENT AND CHARACTERIZATION OF MONOCLONAL ANTIBODIES TO LYMPHOCYTE CELL SURFACE ANTIGENS OF THE RHESUS MONKEY

Three monoclonal antibodies directed against rhesus lymphocyte cell surface antigens are described. A pan-T mAb, T64, and a T suppressor mAb, T35, showed phenotypic and functional specificity for both human and rhesus cells. In contrast, a third mAb, N42, identifying natural killer cells in rhesus peripheral blood leukocytes, was not crossreactive with the corresponding homologous human cells. N42 reacted with the same cells identified by Leu 11a and Leu 11b in rhesus PBL and in a functional assay decreased NK activity by 80%. N42 precipitated a 50KD protein from rhesus PBL lysates and was reactive with the Fc receptor domain of the NK cell. The T-S functional activity of cells reactive with mAb T35 was demonstrated in a pokeweed-mitogen-driven system for Ig synthesis: removal of the T35 positive cells by complement-mediated lysis led to an enhanced production of Ig by rhesus PBL, and the addition of T35 positive cells to a culture of T helper and B cells resulted in a reduction of this response. T35 was determined to be an IgG2a immunoglobulin and precipitated a 34KD protein from rhesus cell lysates. An IgM immunoglobulin, mAb T64 delineated all T lymphocytes, inhibited E-rosette formation, interfered with the proliferation of cells stimulated with mitogens, and precipitated a 52KD membrane protein. The potential utilization of these mAbs in vivo for organ or tissue transplantation in the rhesus monkey is discussed.

Surgical influence on murine immunity and tumor growth: relationship of body temperature and hormones with splenocytes.

Adrenalectomy predisposed the C3HeB/FeJ Mouse to tumor from a low dose of tumor cells, derived from a C3H spontaneous mammary adenocarcinoma. Sham surgery had a similar effect. In contrast, ovariectomized females, intact females, and male mice did not allow the low dose of cells to develop into a tumor. In order to better understand the role of hormones on the immune system controlling tumor growth, normal C3HeB/FeJ mice were studied for the effect of corticosterone or estradiol on splenic lymphocyte surface antigen expression. Adrenalectomy and ovariectomy caused a decrease in the percentage of all T cell subclasses and an increase in absolute numbers of immunoglobulin-bearing cells. Reconstitution of ovariectomized mice with estradiol did not significantly alter lymphocyte cell surface antigen expression. In contrast, injection of corticosterone into adrenalectomized mice brought these values to normal. Further study on normal mice placed on a 12:12-hr light:dark schedule showed that the hours after lights on (HALO) had a significant effect (analysis of variance) on body temperature, percentage of splenic B cells, T pan, T helper and T suppressor cells, and absolute numbers of T pan cells. Brain dehydroepiandrosterone sulfate correlated positively with T pan lymphocytes, but showed no significant effect on HALO. In contrast, body temperature showed a strong circadian rhythm (P less than 0.001). In addition, the presentation of estrus was circadian rhythmic (P = 0.003) with 58% of mice in estrus at 16 HALO and only 8% at 4 HALO. Multiple regression analysis revealed body temperature was strongly associated with absolute numbers of splenic T lymphocytes and their subsets, as well as percentage of B lymphocytes, Thy 1.2-, and Lyt-2-bearing cells. Similarly, HALO and estrous cycle stage were associated with percentage of T helper cells. The data showed that body temperature and hormones were associated with the cell surface antigens on lymphocytes and suggest that they affect lymphocyte function.

A rapid staining procedure for two-color analysis of lymphocyte antigen expression

Two color immunofluorescence analysis of lymphocyte cell surface antigen expression using an unconjugated plus a biotinylated monoclonal antibody (mAb) requires four incubation steps: (1) unconjugated mAb; (2) fluorochrome-labelled goat anti-mouse Ig; (3) biotinylated mAb; (4) fluorochrome-labelled avidin or streptavidin. We describe a time-saving modification of this procedure which requires only two incubation steps: (1) simultaneous unconjugated and biotinylated mAbs; (2) fluorochrome-labelled avidin/streptavidin followed by fluorochrome-labelled goat anti-mouse Ig. The slightly delayed (5 min) addition of the goat anti-mouse Ig prevents it from binding to the mAb which has already interacted with avidin/streptavidin. Both procedures yield identical results with a variety of different mAbs.

Sgp-60, a signal-transducing glycoprotein concerned with T cell activation through the T cell receptor/CD3 complex.

T cell activation depends not only on the expression of a TCR, but also on that of accessory molecules that function in cell-cell adhesion and/or signal transduction. The subject of this report is the biochemical and functional characterization of what appears to be a novel murine lymphocyte cell surface antigen, provisionally termed sgp-60. Extensive, higher-order cross-linking of this glycoprotein with an anti-sgp-60 mAb and a second-step antibody reagent results in the activation of resting CD4+ T cells in the presence of a second signal. Monovalent or bivalent engagement of sgp-60 by the anti-sgp-60 antibody results in profound and direct inhibition of anti-CD3- or Con A-driven T cell activation, whereas alternative T cell activation via the phosphatidylinositol-linked proteins Thy-1 and TAP/Ly-6A is not affected. These findings raise the possibility that the sgp-60 molecule may be specifically involved in signal transduction through the TCR/CD3 complex and thus point to an important physiologic role for this protein in CD4+ T cells.