An attempt to obtain a specific antiserum forin vitro immunological conditioning of bone marrow in acute non-lymphoblastic leukemia

Summary An antiserum was produced in rabbits to acute undifferentiated leukemia (AUL) antigens. Partial absorption of the antiserum allowed the removal of the cytotoxic activity against normal mononuclear cells of peripheral blood and bone marrow, in contrast to blast cells from acute and chronic leukemias, which were killed by the antiserum in a standard NIH cytotoxicity test. These observations suggest that antigens present on blast cells of AUL are also expressed on some acute and chronic leukemic cells and that these antigens could be interpreted as B or leukemia-associated antigens because of the disappearance of all cytotoxic activity of the immune serum after extensive absorption with B cells from chronic lymphoid leukemia. Supported by grant no. 8001963.96 from theConsiglio Nazionale delle Ricerche (CNR), Roma, Italy andStiftung Volkswagenwerk, Bundesrepublik Deutschland.     

Human B-lymphocyte antigens expressed by lymphocytic and myelocytic leukemia cells. I. Detection by rabbit antisera

A previously uncharacterized human B-lymphocyte antigen has been detected by rabbit antisera raised to papain digests of spleen cell membranes. The unabsorbed sera reacted in both cytotoxicity and immunofluorescent tests with normal B lymphocytes and cultured B-cell lines but not with normal T lymphocytes or cultured T-cell lines. The cytotoxicity titers against B cells were as high as 1:32,000, whereas the same sera undiluted were negative against T cells. By immunofluorescent staining 6-14% of unfractionated normal lymphocytes and 48-85% of B-rich lymphocyte preparations were positive. Normal peripheral blood granulocytes, platelets, erythrocytes, and phytohemagglutinin blasts were negative. The antisera reacted with the same high titers against leukemia cells from approximately 70% of the patients with acute lymphocytic leukemia, acute myelocytic leukemia, chronic myelocytic leukemia, and seven of eight cases of chronic lymphocytic leukemia. From absorption studies it appeared that the same antigen was being expressed by leukemia cells and normal B lymphocytes. Using immunofluorescent staining the anti-B-cell antisera were able to detect positive leukemia cells in the bone marrow of patients with advanced leukemia and to monitor the elimination of these cells after chemotherapy. Soluble B-cell antigen was found in the serum of some leukemia and lymphoma patients do but not in normal serum.     

Membrane-bound immunoglobulins on human leukemic cells. Evidence for humoral immune responses of patients to leukemia-associated antigens.

Immunoglobulins were detected on the membranes of human leukemic cells by a microcytotoxicity technique. A significant percentage of lymphocytes from normal donors failed to react with goat antisera to human heavy chain determinants or to lambda-light chains. Lymphocytes from some normal donors, however, did react with antisera to k-light chains. A high percentage (50-90) of cells from some leukemia patients were killed by antisera to light chains and by one or more antisera to heavy chain determinants. Trypsin treatment of leukemic cells resulted in a loss of cytotoxic activity with all immunoglobulin antisera. Reactivity with the k-light chain antiserum was detectable 2 h after trypsinization of chronic myeloid leukemic (CML) cells and 8 h after treatment of acute lymphocytic leukemic (ALL) cells. Reactivity with the antisera to heavy chain determinants and lambda-light chains could not be detected 8 and 48 h after trypsinization of CML and ALL cells, respectively. The cytotoxic activity of the immunoglobulin antisera to heavy chains was abolished by absorption with the specific immunoglobulin used to define the antisera by precipitation. Eluates (pH 3.2) prepared from leukemic cells which reacted by cytotoxicity with the immunoglobulin antisera were shown to contain immunoglobulins of different heavy chain classes. In addition, some of the eluates had cytotoxic antibody activity to human leukemia cells. The specificity of the eluted antibodies is similar to the specificity previously described for cytophilic antibodies from leukemic patients and nonhuman primate antisera to human leukemia cells. The possible in vitro detection and in vivo significance of the eluted non-complement-fixing antibodies is considered.     

Isolation and immunological characterization of a major surface glycoprotein (gp54) preferentially expressed on certain human B cells

A major membrane glycoprotein with mol wt of approximately 54,000 has been isolated from membrane preparations of B-type lymphoid cell lines. Antiserum prepared against the isolated material specifically precipitated this glycoprotein from membranes labeled by surface radioiodination or by metabolic labeling. This antiserum was shown by complement-mediated cytotoxicity assay, membrane immunofluorescent staining, and by quantitative absorption analysis to react preferentially with certain B-lymphoblastoid cell lines, with a minor population of peripheral blood B lymphocytes, and a major population of tonsillar B lymphocytes. Certain B-cell leukemias also expressed the antigen, whereas others did not. Considerable variability was observed among positive B cells in the intensity of fluorescent staining even among the leukemic cells from a single individual. Although T cells, including T cells, were negative by direct immunofluorescent and cytotoxicity assay, evidence for low levels of the antigen on the cells of T cell lines was obtained. The whole specific antiserum and its F(ab')2 fragments stimulated B lymphocytes to proliferate. This proliferation did not produce differentiation to plasma cells and was T-cell independent. The monovalent Fab fragments had no effect. None of these preparations timulated T cells. The possibility that this antigen, termed gp54, may play some role in B-cell activation is discussed.     

Relationships between membrane antigens of human leukemic cells and oncogenic RNA virus structural components

Leukemic cells from all human chronic granulocytic leukemia (CGL) and some acute myelomonocytic leukemia (AMML) donors are lysed by rabbit antisera to a purified glycoprotein of Friend murine leukemia virus (FLV gp71) in a microcytotoxicity assay. These antisera are not cytotoxic to cells from patients with acute myelocytic leukemia (AML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), or to peripheral blood lymphocytes from normal donors. A goat antiserum to gradient purified FLV in addition to reacting with cells from CGL and AMML donors also reacted with cells from AML patients and some ALL donors. However, this antiserum failed to react with cells from CLL patients. Peripheral blood and bone marrow leukocytes prepared from leukemic patients in clinical remission failed to react with antisera to FLV and FLV gp71. Absorption experiments demonstrated that the antigen on CGL cells which is reacting with the antiserum to FLV gp71 is also present on normal human platelets and neutrophils. Similar absorption studies showed that the antigen on AML cells detected by the FLV antiserum is not present on normal leukocytes and platelets and appears to be related to the major internal p30 antigens of mammalian RNA tumor viruses. Another antigenic relationship between oncornaviruses and membrane antigens of human leukemia cells was shown by the ability of FLV antigens to absorb the cytotoxic reactivity of nonhuman primate antisera detecting human leukemia-associated antigens. FLV and FLV gp71 antigens were able to absorb all cytotoxic activity of monkey and chimpanzee antisera to human myeloid leukemia antigens when these antisera were tested with CGL cells. These two approaches to an analysis of cross-reactivity indicate that the antigenic determinant(s) detected by the cytotoxic reactions of the FLV gp71 antiserum with human CGL cells is different from the determinant on FLV gp71 which is responsible for the inhibition of the reactivity of simian antisera with CGL cells. Since the goat and rabbit antisera to FLV and FLV gp71 are able to distinguish AML from CGL cells by direct cytotoxicity testing and absorption, they may be valuable reagents for the serological diagnosis of myeloid leukemia. In addition, since peripheral blood cells from AML and CGL patients in clinical remission were seronegative, the antisera may be valuable as management aids. The data in this report indicates that whatever the mechanism of leukemogenesis is in man, cells from CGL and AML patients possess certain membrane antigens which cross-react with FLV structural components such as p30 and gp71.     

Identification of a p69,71 complex expressed on human T cells sharing determinants with B-type chronic lymphatic leukemic cells

In the course of generating monoclonal antibodies to human thymus-dependent differentiation antigens, we were able to define specificities shared by T cells and by cells from patients with chronic lymphatic leukemia that were not detectable on normal B cells. In particular, one of these antibodies was reactive by indirect immunofluorescence with greater than 95% of the thymocytes and 80--95% of nonadherent sheep erythrocyte-rosetting peripheral blood lymphocytes (PBL), but was unreactive with normal B cells or cell lines derived from PBL by Epstein-Barr virus transformation. However, the leukemic cells from 11 of 14 patients with B-type chronic lymphatic leukemia were found to express detectable concentrations of this surface determinant. The target antigen recognized by this monoclonal antibody was shown by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be a p69,71 complex. Our findings suggest a possible relationship between this antigen and the previously described GIX system in the mouse.     

Poor Mixed Lymphocyte Reaction Stimulatory Capacity of Leukemic B Cells of Chronic Lymphocytic Leukemia Patients Despite the Presence of Ia Antigens

The human Ia-like antigens, selectively expressed on B lymphocytes, are now recognized to be closely associated with, or identical to, the gene products of the major histocompatibility complex responsible for stimulation in the mixed lymphocyte reaction. The leukemic B lymphocytes of patients with chronic lymphocytic leukemia express these antigens very well. In the present study they were readily detected by several techniques utilizing both allo- and heteroantisera. However, the leukemic B cells from most patients were found to be extremely poor stimulating cells in the mixed lymphocyte reaction. This was particularly apparent when comparisons were made on a B-cell basis with isolated normal B lymphocytes.Leukemic cell death, abnormal kinetics of leukemic cell-mediated stimulation, and serum or cellular suppressor factors do not appear to explain these findings. Studies comparing cells from a leukemic patient with those of her HLA identical sibling and results of mixed lymphocyte reactions between normal and leukemic subjects discordant for D-region-associated Ia antigens ruled out genetic explanations for the differences observed. Experiments with normal peripheral blood mononuclear cells depleted of T cells and monocytes exclude the quantitative deficiency of monocytes which is found in the peripheral blood of most leukemic patients as an explanation.The present results with chronic lymphocytic leukemia cells indicate that the mere expression of the Ia-like antigens by cell populations does not render them effective stimulators. The accumulated evidence obtained indicate that abnormalities, particularly of membrane function and metabolism, known to occur in chronic lymphocytic leukemia lymphocytes may be involved in the poor stimulatory capacity of the leukemic B cells.     

Interaction of Herpesvirus ateles and Hepesvirus saimiri with primate lymphocytes. I. Selective adsorption of virus by lymphoid cells

A semiquantitative infectious bioassay, described by Sairenji and Hinuma for the measurement of Epstein-Barr virus (EBV) receptors, was adapted to study the adsorption of the lymphotropic New World herpesvirus ateles (HVA) to established lymphoid lines and lymphocytes of human versus marmoset origin. All marmoset T-cell lines [HVA- or HVS (herpesvirus saimiri)-carrying] had a high adsorbing capacity. EBV-carrying marmoset or human B-cell lines, one EBV-negative human B-lymphoma line, and one human myeloid leukemia-derived line failed to adsorb detectable virus. Two human T-cell derived cell lines of acute lymphatic leukemia origin showed moderate but significant adsorption, and the same was true for unfractionated marmoset lymphocytes. Human peripheral blood did not adsorb, before or after fractionation, whereas normal human thymocytes showed moderate but significant adsorption. It is concluded that HVA receptors are present on human T cells, probably belonging to certain subclasses.     

Characterization of a new monoclonal antibody 6G7 that recognizes a unique antigen on myeloid and lymphoid cells

We generated a monoclonal antibody (mAb) 6G7, which recognizes a 220-kD antigen on selected subpopulations of normal myeloid and lymphoid cells and their malignant counterparts. 6G7 reacts with 90-95% of peripheral blood B cells, 70-80% of CD8(+) cells, 30-35% of CD4(+) cells, 20-40% of monocytes, and 20-40% of CD34(+) cells from bone marrow. 6G7 reacts with leukemic blasts in acute myeloid leukemia (14/16), adult acute lymphoblastic leukemia (ALL) (5/5), pediatric ALL (5/9), chronic lymphocytic leukemia (8/8), follicular lymphoma (7/7), and Burkitt's lymphoma (1/1). Long-term bone marrow culture of 6G7(+/-) cells showed the majority of clonogenic hematopoietic cells were in mAb 6G7 subpopulation. An immunotoxin of 6G7 and ricin A chain was cytotoxic to 6G7(+) leukemia cell lines. mAb 6G7 has potential clinical applications for targeted immunotherapy of both leukemia and lymphoma.     

Combined Studies of Complement Receptor and Surface Immunoglobulin-Bearing Cells and Sheep Erythrocyte Rosette-Forming Cells in Normal and Leukemic Human Lymphocytes

Human lymphocytes from normal peripheral blood, thymus, spleen, thoracic duct, and peripheral lymphocytes from patients with chronic lymphatic leukemia were studied for complement receptor sites (CRL), surface immunoglobulin (SIg), and for the ability to form rosettes with sheep erythrocytes (TRFC). The two B cell markers (CRL and SIg) were found to be in overlapping, but not totally identical populations, whereas cells that were able to form rosettes were found in a totally unrelated population of lymphocytes; TRFC is therefore probably a reliable marker for T cells. In peripheral blood 24% of lymphocytes had SIg, but only half of these were also CRL. Almost all of the non-SIg peripheral blood lymphocytes were TRFC. In the spleen and thoracic duct only a few lymphocytes were observed that had SIg and were not CRL. On the other hand, in two of three spleens studied 10-20% of cells were CRL that did not have SIg. In the thoracic duct all non-CRL that did not have SIg. In the thoracic duct all non-CRL, non-SIg cells were TRFC. In chronic lymphatic leukemia three findings were made: (a) The presence or absence of CRL was independent of the presence or absence of SIg so that in individuals whose cells were non-SIg. CRL were usually plentiful. (b) Leukemic cells were essentially negative for TRFC. (c) Leukemic cells reacted poorly with human C3 compared to mouse C3, EACmo detecting up to 20-fold more CRL than EAChu. This latter finding was in sharp contrast to normal CRL that reacted somewhat preferentially with EAChu. These data suggest that altered surface Ig receptors and complement receptors are present in chronic lymphatic leukemic cells. Since the cells obtained from all leukemic patients tested in this study had either the complement receptor or surface immunoglobulin in a high percentage of their cells and were essentially negative for TRFC, it is strongly suggested that leukemic lymphocytes are of B cell origin. The finding of lymphocytes with only one of the two B cell markers suggests that these markers are not uniformly present on all B cells and that depending on the source, one or the other may be deficient.     

Studies of T- and B-Lymphocytes in Patients with Connective Tissue Diseases

Peripheral blood lymphocytes from normal subjects as well as patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and active tuberculosis were studied for the relative distribution of bone marrow-derived lymphocytes (B-cells) and thymic-derived T-cells. B-cells were identified by direct immunofluorescence of surface Ig markers; T-cells were studied using rabbit antisera to pooled human fetal thymocytes absorbed with chronic lymphatic leukemia lymphocytes as a source of B-cells. In normal subjects, the sum of percentages of peripheral blood lymphocytes staining for surface Ig (B-cells) plus the percentage of cells staining with the absorbed antithymocyte antiserum closely approximated 100%. The mean value for percent B-cells among 51 normals tested was 22.9%±7.1; mean T-cells value was 75.3±13.95%. T-cell-specific antiserum stained 18% of normal human bone marrow lymphocytes, 42.5% of lymphocytes from normal spleens, and 98% of cells obtained from thoracic duct drainage of patients with RA. Specificity of antihuman thymocyte antiserum appeared to depend on the use of living cells.     

Ia-bearing T lymphocytes in man. Their identification and role in the generation of allogeneic helper activity

The presence of Ia-like antigens was demonstrated on a small population (2-6%) of highly purified human circulating T lymphocytes by immunofluorescence with a rabbit anti-Ia serum raised against the isolated bimolecular Ia structure. The Ia+ T lymphocytes have no surface or intracellular immunoglobulins. The expansion of this Ia+ T- cell population was encountered in certain patients. Ia antigens were also found on T blasts grown in long-term cultures with conditioned medium generated by phytohemagglutinin-stimulated lymphocytes. In addition, leukemia blasts which stained for Ia antigens and formed E rosettes were identified in the peripheral blood of two leukemic patients. This evidence further supports the existence of Ia-bearing T cells in man. The Ia+ T-lymphocyte population was shown to contain cells responsible for the generation of allogeneic helper activity. Elimination of Ia+ lymphocytes from a purified T-cell population by the anti-Ia antiserum and complement abolished its ability to help an allogeneic B-cell preparation to generate plaque-forming cells against sheep erythrocytes in vitro in the presence of the antigen.     

Autologous leukemia-specific T-cell-mediated lymphocytotoxicity in patients with acute myelogenous leukemia

Short-term culture of acute myelogenous leukemia patient's remission lymphocytes with inactivated autologous leukemic blast cells plus allogeneic lymphocytes, generated effector T lymphocytes which were cytotoxic for the specific autologous blast cell in 11 of 14 patients studied. Experiments using Daudi and Molt 4 lymphoblastoid cell lines as third-party helper cell suggest that an HLA D locus incompatability is necessary to provide effective help in this system. Cold target inhibition experiments, crossover studies between pairs of patients, and experiments with allogeneic leukemic blast cells as priming stimulus suggest that the target antigen is only present on the specific autologous blast cell.     

Occult leukocyte and tumor-associated antigens assessed by flow cytometry: A Review

We have developed a method for assessing the reactivity of monoclonal antibodies with intracellular antigens. This procedure has been utilized to evaluate the reliability of leukocyte surface antigens as markers of lymphoid cell lineages and for leukemic cell classification, and to further examine the specificity of tumor-associated antigens. Occult expression of certain leukocyte antigens was found in human large granular lymphocytes (LGL), the cells responsible for natural killer (NK) activity. These included the T65 antigen, a T-cell-associated antigen recognized by monoclonal antibodies T101 and Leu-1; the 140Kd 6–cell-associated glycoprotein recognized by monoclonal antibody 82; and the molecules recognized by the monocytel macrophage antibody MO-2. Thus, T-cell, B-cell, and monocyte markers were detected intracellularly in human LGL but not on the surface. Similarly, normal T-cells expressed the 6-2 and MO-2 markers intracellularly. The pan-T-cell antigen Leu-4 (OKT-3) was found to be expressed intracellularly in normal monocytes and in B-cell chronic lymphocytic leukemia cells. In addition, a secreted 100Kd human melanoma-associated antigen as well as vH-ras-p21 oncogene product could be demonstrated intracellularly in high amounts in human tumor and viral-transformed cells. These results indicate that leukocyte differentiation and tumor-associated antigens are expressed in a wider variety of cells than previously detected on the cell surface; and therefore cell lineage and specificity studies require a reassessment.     

Recognition by pregnancy serums of non-HL-A alloantigens selectively expressed on B lymphocytes

A group of alloantibodies are found in pregnancy sera which react with antigens present on B lymphocytes and monocytes but are not detectable on the vast majority of unstimulated T cells. This specificity distinguishes them from HL-A antibodies which react with both cell types. They were readily recognized through indirect fluorescent antibody analysis by employing the combination of B-cell lymphoid lines and normal peripheral blood T cells. Different sera gave a variety of patterns of reactivity with a panel of 11 lymphoid lines. Similar differential patterns were also observed with normal B cells from different individuals particularly after concentrating the B cells. The antibodies were also cytotoxic to B cells and this procedure gave parallel results to the fluorescence method. The pattern of reactions obtained indicated a very heterogeneous system similar to that for HL-A. Special study of certain of the sera provided evidence that the lymphocyte-defined determinants of the mixed lymphocyte reaction system were involved. For convenience the term HL-B has been employed for these antigens.     

Autoantibodies to B Lymphocytes in a Patient with Hypoimmunoglobulinemia: CHARACTERIZATION AND PATHOGENIC ROLE

In a young woman with ulcerative colitis, hypoimmunoglobulinemia, and humoral immunodeficiency, lymphocyte counts vary between 600 and 1,000 per mm(3) with 0.5-1.5% bone marrow-derived (B) cells and 98-99% thymus-derived (T) cells. Anti-lymphocyte antibodies were detected by immunofluorescence and by microlymphocytotoxicity with increased reactivity at +4 degrees C. They belonged to the IgM class and were polyclonal. Studies performed with various normal lymphocyte subpopulations, several lymphoblastoid cell lines and lymphocytes from immunodeficiency patients showed that these antibodies reacted with B cells. The corresponding antigen(s) is distinct from membrane-bound immunoglobulins, is not an alloantigen, and is probably unrelated to the la-like molecules. Pokeweed mitogen stimulated B cells appear to lose this antigen. Cells from various lymphoproliferative disorders were tested. T-derived and "non T-non-B" leukemic cells did not react with the antibody. Malignant cells from B-derived lymphomas and prolymphocytic leukemias were reactive. The incidence of positivity of the leukemic cells among patients with common B chronic lymphocytic leukemia was surprisingly low (one-third of the patients).The autoantibody nature of the anti-B-cell antibodies and their pathogenic role in the genesis of the patient's hypoimmunoglobulinemia was demonstrated by the effect of removal of antibodies by massive plasmaphereses which were followed by a dramatic and transitory increase of B-cell figures. Whereas most primary immunodeficiency syndromes appear to result from an arrest in the differentiation capabilities of immunologically competent cells, autoantibodies to circulating B lymphocytes may be incriminated in the pathogenesis of some cases of hypogammaglobulinemia.     

The common acute lymphoid leukemia antigen on b cells of chronic lymphoid leukemia and non-Hodgkin's lymphoma

Fourteen out of 21 non-Hodgkin lymphoma (NHL) and 3/11 chronic lymphoid leukemia cells (CLL) had the common acute lymphoid leukemia antigen (CALLA) All 32 patients had monoclonal B-cell proliferation. The CLL patients had 90% CALLA positive cells while the proportion of their leukemic elements was superior. Lymph-nodes or bone marrow invaded by a B monoclonal tumor cell population of NHL had significantly more CALLA positive cells (42.1 +/- 32.5%) than non-invaded tissues (11.4 +/- 10.3%). In NHL tissues with monoclonal B-cells, lymph-nodes had significantly more CALLA positive cells (56.0 +/- 29.9%) than marrow (23.5 +/- 27.7%). It is well known that the (CALLA) is not specific for ALL. It has been believed to be a differentiation antigen on pre B-cells. The present study confirms that it also occurs on B-cells (2,4,6,7,8,9,10,11).     

Mechanisms of Lymphocyte Activation: THE ROLE OF SUPPRESSOR CELLS IN THE PROLIFERATIVE RESPONSES OF CHRONIC LYMPHATIC LEUKEMIA LYMPHOCYTES

Binding of (125)I-leukoagglutinin (LPHA) to lymphocyte membrane receptors at equilibrium generated similar curvilinear Scatchard plots in 20 patients with bursa-derived (B)-cell-type chronic lymphatic leukemia (CLL) and 15 controls. If biphasic plots are assumed, the two linear components show markedly diminished receptor capacity (15 and 137 ng/10(6) lymphocytes) in CLL as compared to controls (60 and 668 ng). In contrast, affinity was similar in patients (1.0 x 10(8) M(-1) and 2.1 x 10(6) M(-1)) and controls (1.8 x 10(8) M(-1) and 1.5 x 10(6) M(-1)). Highly purified B cells from patients and controls generated binding data comparable to that obtained from the mixed lymphocyte (ML) suspensions from which they originated. Maximal DNA synthesis of highly purified, normal, thymus-derived (T) and B cells in response to LPHA stimulation was comparable to that of ML (mitotic index [MI] 19.9, 20.1, and 23.4, respectively), though B-cell responses were slightly delayed. In CLL the markedly decreased and delayed DNA synthesis by ML (MI 2.3), and by highly purified T (MI 1.6) and B (MI 1.9) cells seemed out of proportion to their decreased receptor capacity for LPHA. The impaired mitogenic responses of leukemic cells from five patients were not enhanced when cocultured with normal lymphocytes. In contrast, cells from eight patients inhibited cocultured normal lymphocyte responses to LPHA by 94.3%. Sera from these patients and supernates from their cultured cells did not mediate this suppressor effect. These observations indicate that the decreased DNA synthesis observed in CLL is not an attribute of B cells and does not represent the expected response of a few residual normal T lymphocytes, but rather reflects impaired responses by all CLL cells. The defect does not relate to the density or function of membrane receptors for LPHA, to the presence of inhibitors in these patients' sera, or to depletion of helper T cells. Our data strongly suggest that one mechanism for the immunoincompetence observed in CLL reflects excessive suppressor-cell activity.     

Diagnosis of cutaneous T cell lymphoma by use of monoclonal antibodies reactive with tumor-associated antigens.

Two murine monoclonal antibodies (BE1 and BE2), produced by using leukemic helper T cells from a patient with cutaneous T-cell lymphoma (CTCL) as immunogens, reacted selectively with CTCL lymphocytes and some transformed cultured lymphocytes, as determined by radioimmunoassay (RIA) and indirect immunofluorescence (IIF). BE1 reacted significantly (P less than or equal to 0.001) with leukemic CTCL lymphocytes and with CTCL cells from infiltrated lymph nodes (RIA, mean +/- SD = 776 +/- 275 cpm), as compared with background counts (263 +/- 68). BE1 binding to normal blood mononuclear cells (RIA, mean +/- SD = 283 +/- 58 cpm) was indistinguishable from background. BE1 also reacted with Epstein-Barr virus (EBV)-transformed B-cell lines (RIA, mean +/- SD = 794 +/- 230) and some long-term T-cell lines. BE1 did not react with the majority of lymphoid cell lines or tumor cell lines tested. BE1 also did not react with any normal tissues screened by IIF. BE1 precipitated a molecule from CTCL cells that, under reducing conditions, has two components with molecular mass of 27,200 and 25,800 D. BE2 also reacted significantly (P less than or equal to 0.001) with CTCL cells from two of four patients (RIA, mean +/- SD = 519 +/- 113 cpm). The binding of BE2 to normal mononuclear cells was indistinguishable from background (309 +/- 38 cpm). BE2 also reacted with an antigen present on EBV-B-cell lines (RIA, mean +/- SD = 654 +/- 194) and MOLT 3 and HUT 78 T-cell lines. BE2 reacted with an antigen expressed on a subpopulation of lymphocytes from five of eight patients with B-cell CLL studied by IIF (mean +/- SD = 18 +/- 6). Other long-term T-cell lines and tumor cell lines studied by IIF were unreactive with BE2. BE2 did not react with any of the normal tissues studied. BE2 precipitated a molecule (78,000 D) from CTCL cells and EBV-B cells with a single component under reducing conditions. Immunoperoxidase-labeled BE1 and BE2 reacted with CTCL cells in frozen sections of infiltrated lymph nodes and skin. In addition, BE1 and BE2 reacted with blood lymphocytes from 16 of 21 patients whose CTCL had otherwise been considered localized to skin. These two monoclonal antibodies react with tumor antigens associated with CTCL and appear to be useful in the diagnosis of this disorder.     

Unique leukemic non-T/non-B lymphoid cell lines (REH and KM-3): absence of MLR-S and presence of suppressor cell activity for normal T-cell response.

The present study unequivocally demonstrates that leukemic non-T/non-B lymphod cells from three cell lines (NALL-1, NALM-6 and NALM-16) possess a strong stimulating capacity in "one-way" mixed lymphocyte reaction (MLR-S), while leukemic cells from two non-T/non-B cell lines (REH and KM-3) possess no MLR-S. It is speculated that leukemic non-T/non-B lymphoid cells with MLR-S may represent less differentiated leukemic B cells and leukemic non-T/B lymphoid cells without MLR-S may represent less differentiated leukemic T cells. The REH or KM-3 cells without MLR-S also act as suppressor cells on normal T lymphocyte response to mitogen and allogeneic cells by secreting a potent suppressor activity. The MOLT-4 leukemic T lymphoid cells with no MLR-S, on the other hand, do not act as suppressor cells on T lymphocyte response. The soluble factor(s) secreted by the REH or KM-3 cell line is non-toxic to T lymphocytes and heat-sensitive. A significant suppression of T lymphocyte response is still observed, even when the active material is only present for one hour prior to the addition of PHA or it is added several days after the beginning of cultures. The biological and physico-chemical nature of this active material has not been defined. Further studies are currently in progress for biological and physico-chemical characterization and isolation of the active material.