Murine macrophage cell line AP284 presents antigen to cloned MT4+, Lyt-2- T cells in vitro and in vivo

A murine macrophage cell line AP284 that appeared to be mature in phenotype was isolated. After repeated cloning, the cell line expressed the markers Mac-1, Mac-2, Mac-3, 2.4G2, F4/80 as well as Ia antigens. Moreover, it was positive for the enzymes nonspecific esterase and acid phosphatase, negative for alkaline phosphatase and was able to phagocytize latex beads. We studied whether this cell line was able to present antigen to cloned MT4+, Lyt-2- T cells specific for methylated bovine serum albumin (mBSA) or ovalbumin (OVA). The in vitro proliferative response of the cloned T cells specific for mBSA or OVA was found to be effectively supported by AP284. This proliferation could be blocked by monoclonal antibodies against Ia determinants. AP284 also effectively presented antigen in vivo as was shown in a foot swelling assay measuring delayed type hypersensitivity (DTH) to mBSA caused by specific cloned T cells with the helper phenotype. This offers a unique model system for studying the process of antigen presentation in which both the antigen presenting cells and the T cells are monoclonal.     

Different reticular elements in rat lymphoid tissue identified by localization of Ia, Thy-1 and MRC OX 2 antigens.

An indirect immunoperoxidase method was used to localize the Ia, Thy-1 and MRC OX 2 antigens in rat lymphoid tissues. The antigens were detected using mouse monoclonal antibodies and all were notable since they were found to be expressed on different dendritic or reticular elements in lymphoid tissue. Ia antigen was present on bone marrow-derived dendritic cells in the T-dependent areas of spleen and lymph nodes in addition to B cells. MRC OX 2 antibody labelled dendritic cells in the follicles of spleen and lymph nodes but not the Ia positive cells in the T-dependent areas. Cells associated with blood vessels including the endothelium of the post capillary venules were also labelled with MRC OX 2 antibody. In contrast, anti-Thy-1 antibody labelled the pericyte sheath surrounding the post capillary venules and connective tissue elements in lymphoid tissues. The odd patterns of distribution of these antigens are discussed with respect to possible functions of the antigens and the cells they label.     

Nodular sclerosing, mixed cellularity and lymphocyte-depleted variants of Hodgkin''s disease are probable dendritic cell malignancies.

The normal counterpart of the Reed-Sternberg cell and its mononuclear variant, collectively referred to as Hodgkin's cells (HC), remains controversial. The possibility that HC are malignant dendritic cells was tested by using a panel of 38 monoclonal antibodies to phenotype the cells from 16 cases of Hodgkin's disease (HD), excluding lymphocyte-predominant HD, and the Hodgkin's cell line L428. The results were then compared with the known phenotype of human dendritic cells. HC stained strongly for HLA Class I and Class II antigens. The leucocyte common antigen was weakly expressed in most cases. Expression of T and B cell markers was unusual, with the exception of the CD40 antigen which was found on a majority of HC. HC commonly expressed the CD11a, CR4 (CD11c), CD15, CD18 and a number of activation antigens but did not stain with a variety of macrophage-specific antibodies. The antigenic phenotype of L428 and the HC of case material were similar. This immunocytological analysis failed to support a lymphocyte or macrophage origin for HC. Instead the antigenic phenotype of the Reed-Sternberg cell and its mononuclear variant more closely resembles that of dendritic cells than of any other haemopoietic cell normally resident in lymph nodes.     

Tissue localization and retention of antigen in relation to the immune response

Antigen persists for months or even years in lymphoid tissues of immune animals and this antigen is believed to participate in the induction and maintenance of B-cell memory as well as in the maintenance of serum antibody levels. In the present report we describe the phenomenon of antigen localization and long-term retention on mouse follicular dendritic cells (FDCs). The antigens used were injected in the hind footpads of immune mice and the popliteal lymph nodes were the lymphoid organs generally studied. In addition to presenting the morphological features of mouse FDCs, we report the results of a study of the mechanism of antigen migration from the site of initial localization in the lymph node subcapsular sinus to the regions of follicular retention in the cortex. The migration was followed by light and electron microscopy. The results support the concepts that immune complexes are trapped in the subcapsular sinus and are transported by a group of nonphagocytic cells to follicular regions. The mechanism of transport may involve either migration of pre-FDCs with a concomitant maturation into FDCs, or cell-to-cell transport utilizing dendritic cell processes and membrane fluidity; or a combination of the two mechanisms may be in operation.     

The differential diagnosis of hairy cell leukemia with a panel of monoclonal antibodies

A panel of monoclonal antibodies reactive with normal human lymphoid cells and with hairy cells has been applied to the immunocytochemical analysis of hairy cell leukemia. Staining was performed by immunoenzymatic methods on frozen sections of bone marrow trephines and extramedullary tissues and on cell smears. Hairy cells reacted with antibodies against HLA-DR, leukocyte common antigen, B-cell antigens (antibodies To15 and B1) and with three anti-hairy cell monoclonal antibodies (S-HCL3, HC1, and HC2). Neoplastic cells in other B-cell lymphoproliferative disorders also expressed HLA-DR, leukocyte common, and B-cell antigens but were consistently negative for the antigen detected by monoclonal antibody S-HCL3. Furthermore, hairy cells differed from other neoplastic B-cells in that they were unreactive with monoclonal antibodies against C3b receptors, anti-Leu-1, Tü1, Tü33, and lacked a meshwork of dendritic reticulum cells. These findings establish a distinctive antigenic phenotype for hairy cell leukemia and indicate that it may be diagnosed reliably by immunoenzymatic labeling of tissue sections or cell smears.     

Thy-1 positive tingible body macrophages (TBM) in mouse lymph nodes

This study was prompted by the observation that cells in mouse lymph nodes (LN) with cytological characteristics of tingible body macrophages (TBM) appeared to be Thy-1 positive. The objective of this study was to determine if these large cells were TBM and to conclusively demonstrate their reactivity for Thy-1. The cells were studied using monoclonal antibodies (MoAb) against Thy-1 and macrophage markers including F4/80 and Ia antigens at both light microscopic (LM) and electron microscopic (EM) levels. Immunocytochemical reactivity by TBM for Thy-1 antigen specific MoAb was demonstrated by LM in both in situ and in vitro LN preparations. Furthermore, ultrastructural examination of these germinal center cells in situ demonstrated that the Thy-1 reactivity visualized at the LM level was associated with ribosomes and endoplasmic reticulum as well as their plasma membrane. Similarly, these cells expressed intracytoplasmic and membrane reactivity for Ia antigens and also for the macrophage specific antigen F4/80. This indicates that the reactivity is due to active synthesis of the Thy-1 antigen and not attributable to reactivity of any phagocytosed Thy-1 positive cells. As defined by their germinal center location and morphological characteristics, these Thy-1 reactive macrophages were identified as TBM. Germinal center TBM thus represent a unique, vigorously phagocytic subset of mature macrophages which express both macrophage and thymocyte markers.     

Dendritic cell and macrophage staining by monoclonal antibodies in tissue sections and epidermal sheets.

Mouse tissue sections were stained by monoclonal antibodies to macrophage antigens (Mac-1 (M1/70), Mac-2 (M3/38), Mac-3 (M3/84) with the use of immunoperoxidase. Mac-1 was located diffusely in the cytoplasm of round cells in a high percentage of alveolar macrophages, resident peritoneal and bone marrow cells, in splenic red pulp, and in rare perivascular cells in the thymus. Mac-1 was absent in epithelial cells and Langerhans cells. Mac-2 was strongly positive in many dendritic cells in the thymic medulla, more than the cortex, in paracortex and medulla of lymph nodes, sparing the follicles, and in the marginal zone of spleen. There were a few positive cells in germinal centers. Mac-2 was located in a low percentage of bone marrow and a high percentage of resident peritoneal cells. When positive in sections Mac-3 always showed granular cytoplasmic staining. Bone marrow showed a high percentage of cytoplasmic staining (greater than 50%), as compared with low surface staining (less than 1%). It was found in hematopoietic cells, and in all endothelium, including postcapillary venules and lining of sinuses. It was probable that the resulting dendritic staining pattern for Mac-3 in paracortex of lymph node, white and red pulp, thymic cortex, and medulla included dendritic cells other than endothelial cells. Alveolar macrophages and Kupffer cells were positive for Mac-2 and Mac-3. Mac-3 also stained bile canaliculi. Clearly different staining patterns were found in epithelial cells for Mac-2 and Mac-3 in kidney tubules, intestinal mucosal lining, bronchi, choroid plexus, and epidermis.     

Two closely related antigens expressed on granulocytes, macrophages and some reticular elements in rat lymphoid tissues: characterization by monoclonal antibodies

Two distinct novel antigen systems preferentially expressed in rat granulocytes and macrophages were detected using two different monoclonal antibodies (R2-1A6 and R2-2B1). These two antibodies reacted with approximately 50% of rat bone marrow cells, most granulocytes, blood monocytes, alveolar macrophages and peptone-elicited peritoneal macrophages, but not with red blood cells, platelets, thymocytes and T lymphocytes. In addition, R2-2B1 but not R2-1A6 antibody cross-reacted weakly with rat B cells. These two monoclonals also reacted with some reticular elements in rat lymphoid organs including epithelial reticular cells in the thymic medulla and follicular dendritic cells in the lymphoid germinal centre, as well as with the specialized endothelium in the marginal sinuses of the spleen and the post-capillary venules of the lymph node, where lymphocyte recirculation takes place. These antibodies, however, did not label so-called 'dendritic cells' bearing Ia antigens on their cell surfaces, which were found to be located in the thymic medulla, thymus-dependent areas of rat lymphoid tissues and the interstitium of various non-lymphoid organs, suggesting that these dendritic cells, presumably ascribed to those associated with accesory cell function, are separable from the mononuclear phagocyte system in rats by their different reactivities with R2-1A6 and R2-2B1 antibodies.     

Functional characteristics of the veiled cells in afferent lymph from the rat intestine.

Non-lymphoid veiled cells (VC) in the thoracic duct lymph from mesenteric lymphadenectomized rats have been studied by light microscopy, enzyme histochemistry and scanning electron microscopy. These cells arise in the afferent lymph from the intestine. They have been semi-purified and examined for expression of Ia antigens using an indirect immunoperoxidase technique and monoclonal antibodies. Accessory cell function necessary for mitogen-induced blastogenesis in the thoracic duct lymph from these animals has been correlated with the presence of VC by depletion and reconstitution experiments. Similar results were obtained with lymphocyte suspensions from other rat lymphoid organs and they are contrasted with those from studies on mouse lymphoid cells. Antigen presentation in a secondary in vitro lymphoproliferative assay was also depleted from immunized lymph node cells by removal of endogenous VC and can be reconstituted in a dose-dependent fashion with antigen-pulsed VC from afferent intestinal lymph. In contrast, reconstitution of both mitogen-induced blastogenesis and antigen-induced lymphoproliferation with peritoneal exudate cells was poor, while at high multiplicities of added macrophages, such cells were inhibitory. Afferent intestinal lymph VC were found to transport bacteria and bacterial antigen in rats infected with Salmonella typhimurium. The results are discussed in relation to the lineage of the VC in intestinal afferent lymph, their function as accessory cells and their possible physiological role in transporting antigens from the gut to its regional lymph nodes.     

H9/25 monoclonal antibody recognizes a new allospecificity of mouse lymphocyte subpopulations: Strain and tissue distribution

C3H/He-mg mice were immunized with C57 BL/10 (B10) spleen cells and the immune spleen cells were fused with BALB/c myeloma cells (NS1). One of the monoclonal antibodies (H9/25 antibody) produced by the hybrid cells was studied. It reacts with subpopulations of B 10 lymphocytes as well as some lymphoid tumor lines including some of the Abelson virus-induced leukemias. The antigen recognized by H 9/25 antibody is expressed on lymphocytes from all the B 10 congeneic mice tested as well as some other strains of mice. No linkage between genes coding for the antigen and H-2 loci was found as judged by its presence on cells of the B 10 strains regardless of H-2 type and the distribution of the antigen on Bailey recombinant inbred mice. The antigen is expressed on subpopulations of lymph node cells, spleen cells, thymocytes and bone marrow cells. The strain distribution of the H9/25 antigen seems to be identical to that of Ly-6, Ly-8 and Ala-1 antigens. However, the tissue distribution of the antigen recognized by H9/25 antibody, while similar to these alloantigens, is unique and the antigen may be distinct from the other alloantigens.     

Reactivity of monoclonal antibodies against human leucocyte antigens with lymphocytes of non-human primate origin

The phylogenetic distribution of antigens present on human lymphocytes was investigated by incubating human or simian cells with murine anti-human monoclonal antibodies and then determining the level of reactivity with a radiolabelled anti-murine IgG reagent. The monoclonal antibodies used were specific for a T-cell antigen, lymphoid and lymphoid:myeloid antigens, Ia antigens, and beta 2 microglobulin. The cells examined included B- and T-lymphoblastoid cell lines and fresh peripheral blood lymphocytes separated by sheep erythrocyte rosetting into T-cell and non T-cell fractions. Results of these studies showed that the antibodies gave complete cross-reactivity with gorilla and chimpanzee cells while B-cell lines of orangutan origin had lost lymphoid and beta 2 microglobulin markers. Gibbon cells and cells of Old World and New World monkeys reacted strongly only with monoclonal antibodies against Ia antigenic determinants. These Ia antigens were found on the non T-cell fraction of fresh peripheral lymphocytes, on B-cell lines and on some virus induced T-cell tumour lines. Immunoprecipitation analysis using the anti-Ia antibodies showed a degree of molecular diversity on owl monkey and marmoset cells compared to the Ia antigens associated with human cells.     

Diffuse malignant lymphoma with cerebriform nuclei: A B-cell lymphoma studied with monoclonal antibodies

A lymph node biopsy performed on a 55-year-old woman with asymptomatic generalized lymphadenopathy revealed a diffuse, malignant lymphoma composed of small to intermediate-sized lymphocytes with cerebriform-shaped nuclei; electron microscopy confirmed the nuclear complexity. The cerebriform nuclear configuration, coupled with an interfollicular pattern of nodal involvement with encroachment upon residual germinal centers, was presumptive of either mycosis fungoides or a peripheral T-cell lymphoma. Immunologic evaluation, however, indicated that the cerebriform lymphocytes represented a monoclonal B-cell population (IgM-IgD, lambda). Staining with monoclonal antibodies disclosed a phenotype of Ia+, B1+, BA-1+, BA-2+, Leu-1+; the neoplastic cells were unreactive with T-cell, lineage-specific antibodies (anti-Leu-2a, -3a, -4, -5) and with J5 (CALLA). In light of the immunophenotype and the distributional pattern, the cerebriform-shaped lymphocytes may represent an extreme morphologic variant of intermediate lymphocytic lymphoma.     

The rat mixed lymphocyte reaction: roles of a dendritic cell in intestinal lymph and T-cell subsets defined by monoclonal antibodies.

Cells present in the intestinal lymph of rats were obtained in large numbers by removing the mesenteric, portal and caecal lymph nodes and cannulating the thoracic duct 6 weeks later. About 1% of the cells present in the thoracic duct lymph of these mesenteric lymphadenectomized rats had striking dendritic morphology, were strongly Ia+ but labelled weakly with monoclonal antibodies that recognize rat B or T cells. It was found that intestinal lymph was highly enriched for cells that stimulated allogeneic T cells in the mixed lymphocyte reaction (MLR) and cells with stimulator activity co-purified with dendritic cells. Thus, these dendritic cells appear phenotypically and functionally similar to the dendritic cells that have been described in the mouse spleen and rat lymph node. The ability of the intestinal lymph cells to stimulate rat T cells was used to determine which of the two subsets of these cells were the prime responders in the rat MLR. These subsets, defined by monoclonal antibodies, have been shown by previous work to display close functional analogies to the Lyt 2+ and Lyt 2- subsets in the mouse and to the two human T-cell subsets that have been defined by monoclonal antibodies. It was found that the T-cell subset that contains the helper cells for antibody responses proliferated when irradiated, fully allogeneic or semi-allogeneic thoracic duct cells were used as stimulators, but the subset containing suppressor T cells did so only in the fully allogeneic system. Detailed studies showed that in the absence of helper cells in the responder population T cells in the stimulator population of helper phenotype were responsible for proliferation of the suppressor T-cell subset observed in fully allogeneic MLRs. Proliferation of the suppressor T-cell subset could be obtained using semi-allogeneic stimulators, provided that the F1 cells were derived from a source containing dendritic cells but it was shown that, as in the case with fully allogeneic stimulators, the helper T cells in the stimulator population were playing an active role. These results demonstrate that proliferation of the suppressor T-cell subset in the rat MLR is dependent on blastogenic activity provided by the helper T-cell subset and suggest that in some situations this blastogenic activity may arise through the recognition, by the helper cells, of environmental antigens presented on dendritic cells. It has been reported that in the human MLR both T-cell subsets proliferate but that only the helper subset does so when antigen-primed cells are stimulated with specific antigen. The present experiments, by emphasizing the activity of helper T cells in the stimulator population in the MLR, cast doubt on the implication that recognition of alloantigens in vitro differs in an essential way from that of soluble antigens.     

Flow microfluorometric analysis of alloantigen expression during T cell development

The fluorescence-activated cell sorter (FACS) was used to examine the expression of several alloantigens on T cells: Thy-1, Lyt-1.1, Lyt-2.1, Ly-5, Ly-6 and Ly-7. Antibodies secreted by monoclonal cell lines were used to characterize the Thy-1.2, Lyt-1.1 and Lyt-2.1 antigenic determinants, whereas Ly-5.1, Ly-6.2 and Ly-7.2 determinants were identified with alloantisera raised by conventional hyperimmunization. Using indirect immunofluorescence with fluorescein isothiocyanate-conjugated reagents, the profiles obtained with the FACS demonstrated that: (a) the expression of Thy-1 is greater on normal thymocytes than on cortisone-resistant thymocytes (CRT), lymph node and spleen cells; (b) in contrast to Thy-1, the expression of Ly-1 on normal thymocytes is less than on CRT, lymph node and spleen cells; (c) about 90% of normal thymocytes but < 50% of CRT, spleen and lymph node T cells are Lyt-2⁺ and (d) although weakly expressed on normal thymocytes, the amount of Ly-6 and Ly-7 present on peripheral T cells was considerably increased. The significance of these findings and the potential for further analysis of T cell developmental pathways is discussed.     

Tumor immunity meets autoimmunity: antigen levels and dendritic cell maturation

Nai;ve T cells in the draining lymph node (DLN) do not immediately respond to antigenic tissues or antigenic cancers in the periphery. Rather, the conditions under which nai;ve T cells encounter antigen in the DLN can result in the distinct immunological states of ignorance, tolerance or immunity. Recent work suggests that these immunological states are determined by the level of antigen expressed by peripheral tissues and the maturation stage of the dendritic cell presenting the antigen. When antigens are expressed at levels that are sufficient to be cross-presented by mature dendritic cells in the DLN, nai;ve T cells can respond to self antigens or tumor antigens to induce a state of autoimmunity or tumor immunity, respectively. Exploiting these conditions to target unique tumor antigens will enable us to develop better cancer immunotherapies.     

Isolation of functionally active murine follicular dendritic cells

Biochemical, genetic, and immunological studies of follicular dendritic cells (FDCs) have been hampered by difficulty in obtaining adequate numbers of purified cells in a functional state. To address this obstacle, we enriched FDCs by irradiating mice to destroy most lymphocytes, excised the lymph nodes, and gently digested the nodes with an enzyme cocktail to form single cell suspensions. The FDCs in suspension were selected using the specific mAb FDC-M1 with magnetic cell separation technology. We were able to get nearly a million viable lymph node FDCs per mouse at about 90% purity. When examined under light and transmission electron microscopy, the cytological features were characteristic of FDCs. Furthermore, the cells were able to trap and retain immune complexes and were positive for important phenotypic markers including FDC-M1, CD21/35, CD32, CD40, and CD54. Moreover, the purified FDCs exhibited classical FDC accessory activities including: the ability to co-stimulate B cell proliferation, augment antibody responses induced by mitogens or antigens, maintain B cell viability for weeks, and protect B lymphocytes from anti-FAS induced apoptosis. In short, this combination of methods made it possible to obtain a substantial number of highly enriched functional murine FDCs.     

Immunohistologic analysis of lymphoid cells by a rapid double staining method

The development of monoclonal antibodies has allowed characterization of subpopulations of lymphoid cells and of their in situ distribution in tissues. The feasibility of simultaneous localization of two surface antigens was studied by double staining with monoclonal antibodies to B cells, T cell subsets and follicular dendritic reticulum cells (DRC) using the avidin-biotin-complex (ABC) method in sections of human lymphoid tissues (tonsil, lymph node, spleen) and a non-lymphoid tissue, endometrium. Color mixture was avoided when an additional incubation with avidin-biotin-labeled peroxidase and subsequent development in the respective substrate of the first sequence preceded the second staining sequence using the primary antibodies at optimal concentrations. The antigenic profiles were portrayed by contrasting and distinct colors of the reaction products. It was observed that T lymphocytes of the cytotoxic/suppressor and helper/inducer phenotypes were topographically associated with each other and with B cells in B and T cell areas of lymphoid tissues as well as in lymphocytic aggregates in endometria. Subpopulations of these cells were mantled by processes of DRCs in lymphoid follicles. The findings indicate that the double ABC staining method can be used for simultaneous demonstration of two surface antigens in tissue sections.     

Non-activated guinea-pig T cells and thymocytes express Ia antigens: FACS analysis with alloantibodies and monoclonal antibodies.

Conventional alloantisera and monoclonal antibodies to guinea-pig Ia antigens were used for analysis of Ia expression by guinea-pig T cells and thymocytes. Indirect immunofluorescent staining was performed with alloantisera or with ascitic fluid as a source of monoclonal antibody followed by flow microfluorometry analysis on the fluorescence activated cell sorter. About 80% of normal, non-activated peritoneal exudate T cells, lymph node T cells and thymocytes expressed Ia antigens. These data are therefore in contrast to studies with human or murine T cells where Ia antigens were shown to be expressed predominantly on activated but not on non-activated T cells. All the reactivity of the anti-Ia alloantisera for strain 2 T cells could be removed by absorption with an Ia-bearing B cell leukaemia, EN-L2C, but not by its Ia-negative variant, BZ-L2C. Thus, the Ia determinants identified on T and B cells are probably identical. One monoclonal antibody, 25E3, which had previously been shown by serologic analysis to react exclusively with an alloantigenic determinant of strain 2 Ia antigens displayed an unusual pattern of reactivity in that it clearly stained strain 13 thymocytes, but not mature strain 13 T or B lymphocytes. The significance of this possible expression of inappropriate Ia determinants by thymocytes remains unclear. This phenomenon might be associated with differentiation processes in the thymus.     

Phenotypes of immunocompetent cells and Ia antigen expression in oral mucosa and skin of autoimmune mouse strains

The phenotypic characteristics of resident and infiltrating cells in skin and oral mucosa of MRL/Mp-lpr/lpr (MRL-lpr), MRL/Mp+/+ (MRL-(+)), NZB x NZW F1 (NZB/W) and Balb/c mice have been studied using monoclonal antibodies (Mabs) and immunoperoxidase staining with the Avidin-Biotin Complex method. Macroscopically evident skin lesions appeared spontaneously exclusively in aging MRL-lpr mice. Infiltration of lymphocytes was detected within the mucosa and the skin of MRL-lpr-mice. These lymphocytes expressed predominantly L3T4 and to a lesser extent Lyt-2 phenotype (T helper/inducer and T suppressor/cytotoxic subset, respectively). Class II major histocompatibility complex antigen (Ia) expression was detected on macrophages, dendritic cells and few lymphoid cells in normal oral mucosa and skin of all strains examined. However, in the oral mucosa and skin lesions of the 4 months old MRL-lpr mouse also keratinocytes expressed Ia antigens. Keratinocytic Ia expression was also detected in oral mucosa of 8 months old MRL-(+) mice. The described immunopathological findings in skin and oral mucosa of old MRL-lpr mice show a number of important similarities to human SLE disease.     

Studies of murine lymphocyte alloantigens Ly-7.2

The Ly-7.2 alloantigen has previously been defined by an alloantiserum and shown to be expressed on T and B cells; it is now studied for its distribution on responder and stimulator cells in the mixed lymphocyte reaction, on cells responding to mitogens and on mitogen-induced blast cells. In the mixed lymphocyte reaction, the responder cells were Ly-7.2+; stimulator cells were Ly-7.2-. The Ly-7.2 antigen also had an unusual distribution on cells responding to mitogens: leucoagglutinin-responsive cells were Ly-7.2-, concanavalin A Ly-7.2+, pokeweed mitogen Ly-7.2+/-, and lipopolysaccharide Ly-7.2-, i.e., Ly-7 can distinguish between subpopulations of T cells which respond to mitogens (phytohaemagglutinin and concanavalin A). Ly-7.2 was present on approximately 60% of blast cells induced by all mitogens indicating that both Ly-7.2+ and Ly-7.2- cells could be activated to become Ly-7.2+. Further characterization of concanavalin A and leucoagglutinin-stimulated Ly-7.2+ and Ly-7.2- T blast cells with anti-Ly-2.1 and anti-Ia. 17 monoclonal antibodies demonstrated the presence of several T lymphocyte subsets as both Ly-7+ and Ly-7- blast cells contained Ly-2+, Ly-2-, Ia+ and Ia- cells. Ly-7.2 therefore has a heterogenous distribution on normal and activated T and B cell subpopulations, and is a potentially important antigenic marker for studies of lymphocyte differentiation and function.