Immunochemical characterization of human TL-like (T48) and Ly 1-like (T72) glycoproteins using two-dimensional polyacrylamide gel electrophoresis

Xenoantisera, designated AT48 and AT72, were developed by immunizing rabbits with human thymus cell membrane and guinea-pigs with a T-cell glycoprotein purified from leukaemic T-cell membrane. Whereas AT48, after appropriate absorption, reacted exclusively with the majority of thymocytes (mainly cortical thymocytes) among normal lymphoid populations, AT72 reacted with virtually all of the thymus and T cells but not with B cells. Thymocytes, which were strongly reactive with AT72, existed in the thymic medulla, but cortical cells were also very weakly reactive with AT72. When cultured T-cell lines, all of which were derived from patients with T-cell-type acute lymphatic leukaemias, were tested for their reactivities with AT48 and AT72 by immunofluorescence, we found that AT48 stained certain T-cell lines, whereas AT72 stained all of the T-cell lines tested so far. Immunochemical data showed that AT48 precipitated a 48K molecular weight (mol. wt) glycoprotein from 125I-labelled thymus cell surface glycoproteins, which appeared to be very weakly associated with a 12K mol. wt component. These 48K and 12K mol. wt components precipitated by AT48 showed almost identical isoelectric points (pI) to those of HLA heavy chain and beta 2-microglobulin respectively. AT72, on the other hand, precipitated a 72K mol. wt glycoprotein from thymus and T cells as well as from leukaemic T cells. A less prominent 65K mol. wt glycoprotein was also precipitated by AT72 from thymus and T cells but not from leukaemic T cells. These two components showed almost identical pI ranging approximately from 4 to 7, and this marked charge heterogeneity observed was reduced by neuraminidase treatment, suggesting that it reflects the heterogeneity in sialylation of this molecular species. We concluded from these data that AT48 and AT72 used in this work could detect human homologues of mouse TL and Ly 1 antigens respectively.     

Identification of Ia glycoproteins in rat thymus and purification from rat spleen.

A fraction of la-like glycoproteins was prepared from rat thymocytes by lentil lectin affinity chromatography and gel filtration in deoxycholate. Spleen cells from mice immunized with this preparation were fused with myeloma cells to produce antibody-secreting hybrid cell lines. Antibody from four lines called MRC OX, 3, 4, 5, 6 reacted with the la-like glycoproteins, and MRC OX 3 antibody recognized an antigenie determinant polymorphic in the rat. All four antibodies also bound to mouse spleen cells and all detected polymorphisms. Studies on recombinant mouse strains suggest that the determinants are coded by the I-A subregion of the H-2 complex. MRC OX 3 correlates with Ia specificity 9, while MRC OX 4, 5, 6 correlate with specificity 17 or 18. MRC OX 4 monoclonal antibody was used for affinity chromatography to purify Ia glycoproteins from rat spleen. The rat Ia glycoprotein complex was composed of two noncovalently linked polypeptide chains of apparent mol. wt. (unreduced) 30 000 and 24 000 as determined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. The purified Ia glycoprotein partially inhibited the binding to thoracic duct lymphocytes of an alloantiserum which detects Ia antigens linked to the major histocompatibility complex. The monoclonal anti-la antibodies bound to the majority of peripheral B lymphocytes and 18% of thymocytes, but did not significantly bind to peripheral T lymphocytes. There were on average 150000 molecules of Ia glycoprotein per la-positive B lymphocyte, and 45 000 molecules per la-positive thymocyte. From the same fusion, another cell line was prepared called MRC OX 2 which secretes monoclonal antibody to a previously undefined thymus glycoprotein of apparent mol. wt. 60000. Preliminary studies showed that the antigen was expressed on all thymocytes and on peripheral B lymphocytes in smaller amounts. It was also present in brain, but not liver or kidney homogenate.     

Monoclonal antibodies to the murine Ly-2.1 cell surface antigen.

Six monoclonal anti-Ly-2.1 antibodies are described that arose from a single fusion using the spleen from a 129/ReJ mouse immunized with CBA/H thymus cells. The specificity was determined from the strain distribution, and testing of congenic strains where all six monoclonal antibodies detected the Ly-2.1 alloantigen. Furthermore, the antibodies fall into two groups: Group I (four antibodies) detected the expected number of Ly-2+ thymocytes and peripheral T cells, whereas Group II (two antibodies) detected 10% fewer peripheral T cells. Functional studies demonstrated that the Ly-2.1 determinant detected by a Group I antibody (49-31.1) was presented on cytotoxic T cells (Ly-1+2+), on concanavalin A (Con A)-induced suppressor T cells (Ly-1-2+), but absent from helper T cells (Ly-1+2-). One antibody (49-11.1) precipitated a 68,000-75,000 mol.wt glycoprotein, which on reduction yielded 30,000 and 35,000 mol.wt moieties. Thus, by functional, genetic and biochemical criteria these antibodies detected the Ly-2.1 specificity. In addition, a monoclonal, noncytotoxic, IgGl, anti-Thy-1.2 antibody is described.     

Two distinct antigenic markers for rat thymus and T cells defined by monoclonal antibodies.

Rat thymus and T-cell antigens were defined by using two distinct monoclonal antibodies (R-1-3B3 and R-1-12C5). R-1-3B3 antibody, when tested for its reactivity with rat lymphoid cells by immunofluorescence, labelled virtually all thymus and T cells but not B cells and bone marrow cells. The antigen defined by this R-1-3B3 antibody occurred more abundantly on medullary thymocytes and peripheral T cells than on cortical thymocytes. Immunochemical data showed that R-1-3B3 antibody recognized a single glycoprotein with a molecular weight of 67,000, which were able to interact with Lens culinaris haemagglutinin. R-1-12C5 antibody, on the other hand, reacted with all of thymus and T cells as well as with a subpopulation (approximately 20%) of bone marrow cells. In contrast to the antigen defined by R-1-3B3, that detected by R-1-12C5 antibody existed largely on cortical thymocytes and to a much lesser extent on medullary thymocytes and peripheral T cells. R-1-12C5 antibody detected a single glycoprotein with a 95,000 molecular weight, which could also interact with Lens culinaris haemagglutinin. Based on these data described above and since both antigens defined by R-1-3B3 and R-1-12C5 antibodies were absent from rat brain tissue, we concluded that they were distinct from brain-associated thymic antigens in rats including Thy-1 and W3/13 antigen systems.     

Quantitative analysis of tissue distribution of Ly10-like murine alloantigen(s) with antisera and monoclonal antibodies

Tissue distribution of non-Lyt1.1 ("Ly10-like") antigen or antigens encoded by short chromosomal segment differentiating B6-Ly-1a congenic strain from B6 strain of mice was studied by quantitative absorption of (BALB/c X B6)F1 anti B6-Ly-1a antiserum and by direct cytotoxicity of Ly-10-132-12-26 monoclonal antibody on lymphoid cell populations. Identical strain but not tissue distribution pattern does not allow to conclude whether antiserum and monoclonal antibody detect the same or closely linked antigens. Absorption experiments revealed the highest antigen content in the brain tissue, lower in testis and kidney, still lower in lymphoid organs and the lowest in liver and lung. Among lymphoid cells, bone marrow cells had highest absorbing capacity, followed by thymus, spleen and lymph nodes. Monoclonal antibody lysed almost 100% of thymocytes, 30% bone marrow cells and 10-20% of spleen and lymph node cells (both T-cell and B-cell enriched populations contained the same proportion of positive cells). Cortisone resistant thymocytes showed the same sensitivity as cortical thymocytes to Ly-10-132-12-26 antibody which is distinguishable characteristics of medullary thymocytes from peripheral T cells. Mitogen activated lymphocytes exhibited significantly higher expression of Ly10-like antigen than resting peripheral lymphocytes.     

Molecular identification of T cell-specific antigens on human T lymphocytes and thymocytes

We have identified membrane glycoproteins which carry T cell-specific antigens on human T lymphocytes and thymocytes. Purified cells were surface-labeled with NaB³H₄ after treatment with neuraminidase and galactose oxidase. Immunoprecipitations were performed with rabbit anti-human T cell-specific antibodies using co precipitation with protein A-containing staphylococci strain Cowan I. The labeled membrane glycoproteins and the precipitates were subjected to polyacrylamide slab gel electrophoresis and visualized by fluorography. The antibodies specifically precipitated 4 proteins called GP200, GP180, GP165 and GP160 (mol. wts. = 200000, 180000, 165000 and 160000) from surface-labeled T lymphocytes and low-density (medullary) thymocytes. The GP200 and GP180 were not labeled on high-density (cortical) thymocytes. A protein with a mol. wt. of 45000 was precipitated from thymocytes. Another glycoprotein on T lymphocytes and thymocytes with a mol. wt. similar to that of mouse and rat Thy-1 or Θ antigen (mol. wt. 25000) reacted with the antibodies.     

Two distinct antigens on chicken thymocytes defined by monoclonal antibodies

Two distinct antigens expressed on chicken thymocytes were defined by monoclonal antibodies designated as Lc-1 and Lc-2. Lc-1 (IgGl) reacted with 80% of the thymocytes (mostly cortical and medullary thymocytes) and Lc-2 (IgM) reacted with 40% of the thymocytes (mainly cortical thymocytes). Lc-1 reacted with 1% of the spleen lymphocytes, but the antibody was nearly nonreactive with cells from peripheral blood leukocytes, bursa and bone marrow. Lc-2 reacted with only small percentages of spleen and bursa cells, and with very few cells from peripheral blood leukocytes and bone marrow. This antibody reacted with a portion of concanavalin A stimulated spleen lymphocytes. When Marek's disease-derived T-lymphoblastoid cell lines were tested for their reactivities with monoclonal antibodies, Lc-1 reacted with none of the cell lines tested, whereas Lc-2 reacted with four of the six cell lines tested. Antigens recognised by Lc-1 and Lc-2 were first found in chick embryonic thymus on day 13 of incubation, after which the number of cells positive for Lc-1 and Lc-2 rapidly increased, reaching young adult levels by days 15 and 14 of embryonic life, respectively. Lc-1 precipitated materials with apparent molecular weights of 60 and 120 kDa from radioiodinated thymocytes.     

The predominant heavily glycosylated glycoproteins at the surface of rat lymphoid cells are differentiation antigens.

Rat lymphoid cells have been labeled with sodium 3H-borohydride after periodate oxidation. The labeled glycoproteins were solubilized in detergent and analyzed by fluorography after polyacrylamide gel electrophoresis in sodium dodecyl sulfate. Major bands were found at 150 000, 95 000 and 25 000 apparent mol.wt. for thymocytes; at 170 000 and 95 000 mol. wt. for T lymphocytes and at 200 000 mol.wt. for B lymphocytes. Bone marrow cells showed a diffuse band at 100 000 mol.wt. with relatively minor bands around 150 000 mol.wt. With the exception of the 95 000 mol. wt. bands, all these glycoproteins bound to lentil lectin. Using monoclonal or monospecific antibodies in immunoprecipitation and on antibody affinity columns, each of these glycoprotein bands was identified as a previously defined lymphocyte differentiation antigen. The bands at 150 000 mol.wt. on thymocytes, at 170 000 on T lymphocytes, at 200 000 on B lymphocytes, and at 130 000 to 150 000 on bone marrow cells all consist of a leukocyte-common antigen, which has previously been shown to be present on leukocytes but not on other tissues. At least a part of the 95 000 mol.wt. band on thymocytes, T lymphocytes and bone marrow cells is the W3/13 antigen previously shown to be on mature T lymphocytes, polymorphonuclear cells, and in brain. The 25 000 mol.wt. band of thymocytes is the Thy-1 antigen. Similar experiments were carried out on thymocytes labeled with ¹²⁵I by the lactoperoxidase method. An intense band at 150 000 mol. wt. was identified as the leukocyte-common antigen by immunoprecipitation. A labeled band, which did not bind to lentil lectin, was immunoprecipitated at 95 000 mol. wt. with W3/13 antibody. Rat Thy-1 antigen was not labeled with ¹²⁵I.     

Characterization of mouse thymocyte and peripheral lymphocyte xenoantigens by two-dimensional electrophoresis

Two-dimensional electrophoresis was used to analyze the surface glycoproteins of murine thymocytes and lymph node cells. Two-dimensional maps of unselected, radioiodinated lymphocyte surface proteins were complex, showing at least 20 different components, but simpler patterns were obtained by using rabbit antibodies directed against the surface proteins of a T lymphoma cell line, to precipitate xenoantigens from lysates of radioiodinated or biosynthetically labeled thymocytes and lymph node cells. These xenoantibodies precipitated 12–13 distinct components from each cell type, of which all but 3 were sialoglycoproteins. Two types of difference between the surface glycoproteins of thymocytes and peripheral lymphocytes could be detected. First, higher mol. wt. glycoproteins and Thy-1 are more acidic in peripheral lymphocytes than in thymocytes, and this difference disappears after neuraminidase treatment. One additional high mol. wt. glycoprotein is also detectable in peripheral lymphocytes, probably reflecting the greater carbohydrate complexity of these molecules, when expressed on such cells. Second, 3 glycoproteins are strongly labeled only on thymocytes, and 3 others only on peripheral lymphocytes. These 6 glycoproteins might represent genuine differentiation antigens.     

Monoclonal antibody to a human brain-granulocyte-T lymphocyte antigen probably homologous to the W 3/13 antigen of the rat

The monoclonal antibody (F10–44–2) described in this report recognizes an antigen which by quantitative absorption analysis is found predominantly on spleen, lymph node, bone marrow, thymus, granulocytes and brain, the amount of antigen on these tissues being approximately the same within a factor of 2 or 3. Analysis with the fluorescence-activated cell sorter showed that 29% of thymus cells, 61% of bone marrow cells, 95% of blood mononuclear cells, 98% of lymph node lymphocytes and 100% of granulocytes carried the antigen. With blood mononuclear cells and lymph node lymphocytes, there were two distinct peaks, with one peak labeling very weakly. Double labeling experiments established that the weakly labeled peak contained the B lymphocytes. Studies on frozen sections of thymus established that positive thymocytes were found only in the medulla indicating that the antigen appears late in T lymphocyte maturation. The lymphatic nodules (B lymphocyte areas) of spleen and lymph node appeared virtually negative on frozen sections showing that there was too little antigen on the B lymphocyte surface for confident detection by fluorescence microscopy. Sodium dodecyl sulfate polyacrylamide gel eletrophoresis of NaB³ H₄-labeled blood mononuclear cells established that the antigen was a major glycoprotein of the leukocyte membrane and that its mol. wt. was 105 000. This antigen shows a striking similarity in biochemistry and tissue distribution to the W3/13 antigen of the rat and is likely to be the human homologue of this antigen.     

MLR3 molecule is an activation antigen shared by human B, T lymphocytes and T cell precursors

MLR3 molecule is a membrane glycoprotein (mol. mass range 28-34 kDa) present on activated, but not resting human peripheral T cells, B cells and thymocytes. Its kinetics of appearance on the cell surface (3 h after the addition of the inductive signal to the cells) suggests that it is an early activation antigen. The proliferative response of cultured T and B lymphocytes and thymocytes to different activation signals is inhibited by the addition of MLR3 monoclonal antibody. Moreover the antibody in combination with non-mitogenic doses of phorbol myristate acetate leads to proliferation of thymocytes and resting B and T lymphocytes. In the latter, synthesis of interleukin 2 is also induced. Biochemical analysis of MLR3 antigen indicates that it is a phosphorylated protein with N-linked sugar moieties. Together these data suggest a role for MLR3 antigen in the signal transduction process during activation, both for mature lymphocytes and for T cell precursors.     

Analysis of recent thymic emigrants with subset- and maturity-related markers

The thymus plays an integral role in the development and production of T lymphocytes. However, thymocytes differ markedly in their phenotypic characteristics from the T cells normally found in the peripheral lymphoid organs. We have examined the phenotypic characteristics of recent thymic emigrants and compared them with both mature phenotype thymocytes (CD4+ CD8-CD3+ and CD4-CD8+ CD3+) and lymph node T cells. Recent thymic emigrants were defined as those fluorescein-positive cells found in the lymph node up to 16 h after intrathymic injection of fluorescein. Most cells emigrating from the thymus expressed CD3 and either CD4 or CD8, indicating maturity. Recent thymic emigrants, like mature phenotype thymocytes, were slightly larger on average than peripheral T cells, but this differential was lost within 24 h of emigration. Also like mature thymocytes but unlike peripheral T cells, some recent emigrants expressed heat-stable antigen. This did not change within 24 h of emigration. The antigen CD44 (Pgp-1, Ly-24) was expressed on a proportion of mature thymocytes, recent thymic emigrants, and peripheral T cells, and its expression did not show any clear relationship to maturity. The antigen CD45R also did not show marked changes associated with maturity, but our data do not parallel the published data of the expression of CD45R in the human. We conclude that recent thymic emigrants are phenotypically mature with respect to some antigens but not others. None of the antigens we investigated could have been used to uniquely distinguish recent thymic emigrants from peripheral T cells or from mature thymocytes.     

An attempt to produce “pre-T” cell hybridomas and to identify their antigens

With the aim of identifying some of the stages in the development of pre-T cells (cells of the T cell lineage before they enter the thymus), we have produced a large number of hybridomas by the fusion of BALB/c bone marrow cells, bone marrow cells from BALB/c-nu/nu mice, BALB/c fetal liver cells and BALB/c fetal thymocytes with the AKR thymoma BW5147. The hybridomas were selected for the expression of the Thy-1.2 antigen of th3 normal celldonor and for their ability t9produce interleukin 2 (IL 2) upon co-culture with irradiated normal spleen cells. A set of these hybridomas is described in this communication. The hybridomas were then used to immunize rats and to generate monoclonal antibody-producing B cell hybridomas. Most, if not all, of the immunizing hybridomas were derived from pre-T cells as evidenced by the fact that they produce IL 2, and express some of the T cell markers (the Thy-1.2, Ly-1, Ly-2 or L3T4 antigens). The monoclonal antibodies were tested on a panel of pre-T cell hybridomas and on normal cells obtained from spleen, lymph nodes, thymus and bone marrow. The testing was carried out by the microcytotoxicity assay and flow cytometric analysis. Three groups of antibodies could be distinguished. Some antibodies were broadly reactive, being positive with virtually all the clones in the pre-T cell panel and with a substantial fraction of normal lymphoid cells. The identity of the antigens detected by these antibodies remains unknown but they do not seem to correspond to any of the known cell surface markers. Other antibodies reacted only with some of the pre-T cell clones and did not react at all with normal lymphoid cells obtained from adult animals. Finally, other antibodies still reacted only with a minor subpopulation of thymocytes or of thymocytes and bone marrow cells, as well as some of the pre-T cell clones; they did not react with spleen and lymph node cells. These antibodies might be specific for cells in the prethymic phase of the T cell differentiation pathway. They should prove useful for the identification of pre-T cell markers and hence for the isolation of pre-T cells and their functional analysis.     

Mouse monoclonal antibodies against rat major histocompatibility antigens. Two Ia antigens and expression of Ia and class I antigens in rat thymus

A rat Ia (RT-1B) antigen (called Ia-A) equivalent to the mouse I-A product has been defined with mouse monoclonal antibodies (W. R. McMaster and A. F. Williams, Eur. J. Immunol. 1979. 9: 426). To identify other Ia antigens mouse monoclonal antibodies were raised against rat spleen glycoproteins depleted of the Ia-A antigen. An IgG antibody (called MRC OX17) was obtained and used to purify a molecule which had a similar structure to the Ia-A antigen and reacted with anti-Ia alloantibodies. There was no cross-reaction between the two Ia glycoproteins in assays with mouse monoclonal antibodies, alloantibodies or rabbit antibodies. In one alloantiserum almost all the detectable anti-Ia antibodies reacted with a mixture of the two Ia glycoproteins. The MRC OX17 antibody did not bind to mouse cells, but rabbit antibodies to the pure rat glycoprotein cross-reacted and recognized determinants mapping to the mouse I-E region. In the thymus the rat Ia-E antigen was on cortical epithelial and medullary reticular cells. An IgG monoclonal antibody (MRC OX18) to isotypic determinants of rat histocompatibility RT-1A antigens was also produced and used to analyze these antigens on thymus cells. The heavily labeled thymocytes were those with characteristics of mature T lymphocytes. Cortical epithelial cells and medullary dendritic-like cells were also RT-1A positive.     

Lymphocyte cell surface glycoproteins which bind to soybean and peanut lectins.

In cellular immunology, peanut (Arachis hypogaea) lectin has been used to selectively agglutinate immature lymphoid cells and soybean (Glycine max-lectin to agglutinate B lymphocytes. We have used affinity chromatography to study the surface glycoproteins of rat and mouse lymphoid cells which bind to these lectins. Thymocyte and T and B lymphocyte glycoproteins were analysed either without modification (native) or after the removal of sialic acid with neuraminidase (asialo). The only native glycoprotein which was seen to bind to peanut lectin was the 95,000 mol. wt sialoglycoprotein from thymocytes. The equivalent molecules from T lymphocytes bound to peanut lectin only after neuraminidase digestion. Thus the selective agglutination of thymocytes by peanut lectin would seem to be due to a partial lack of sialic acid residues on the O-glycosidically-linked oligosaccharides of the thymocyte sialoglycoprotein. The B lymphocyte form of the leucocyte-common antigen was the only prominent native glycoprotein which was seen to bind to soybean lectin and this probably accounts for the specific binding of this lectin to B cells. The leucocyte-common antigens, in their asialo forms, from thymocytes and B and T lymphocytes differed in their binding to the lectins and this establishes that these glycoproteins which share antigenic determinants differ in their carbohydrate structures.     

Monoclonal antibody (WT 1) directed against a T cell surface glycoprotein: characteristics and immunosuppressive activity

WT 1, an IgG2a subclass monoclonal antibody, recognizes a human T lineage specific antigen (mol. wt 40,000). This antigen is strongly expressed on thymic T blasts, and on peripheral T cells activated by phytohaemagglutinin, whereas cortical thymocytes and peripheral T cells are moderately positive for WT 1. In contrast, B lymphocytes, myeloid and erythroid cells, including the progenitor cells of these lineages, and terminal deoxynucleotidyl transferase positive cells in the bone marrow, are all WT 1 negative. Binding of WT 1 to T cells is blocked by a previously described antibody (3A1) suggesting that both antibodies bind to the same antigen present on human T cells. WT 1, however, is also reactive with T lymphocytes from rhesus monkeys whereas 3A1 is not. Therefore, the biological effects of WT 1 could be studied in a monkey model. In a skin allograft model, WT 1 was immunosuppressive and induced a marked prolongation of graft survival.     

T Lymphocytes Displaying Major Histocompatibility Complex-Unrestricted Cytotoxicity after Activation by K46M, a Mitogenic Monoclonal Antibody against Leucoagglutinin-Reactive Human T Lymphocyte Surface Components

Activation of human peripheral blood lymphocytes (PBL) with the mitogenic monoclonal antibody (MoAb) K46M, which recognizes 1-5% of PBL, resulted in the expansion of cells with cytolytic activity. Thus, after culture of the activated lymphocytes in medium containing interleukin 2 (IL-2), they lysed a variety of cultured cell lines. The majority of the activated lymphocytes reacted with MoAb to CD8, CD3, and to the T cell antigen receptor heterodimer (Ti) but not with antibodies to antigens expressed on natural killer (NK) cells. The cytotoxicity was not inhibited by MoAb to CD3 or Ti. However, the killing of K562, but not of other cell lines, was enhanced by three to four times in the presence of anti-Ti antibodies. Anti-CD3 or other control antibodies had no effect. Cold target inhibition experiments indicated that the cytolytic lymphocytes recognized closely related structures on the target cells. Phenotypically and functionally similar effector cells emerged after activation of PBL with the anti-CD3 MoAb OKT3. Taken together, the results indicate that activation of PBL with MoAb K46M induces cytotoxic cells that differ from classical NK cells but that resemble mature cytotoxic T lymphocytes (CTL). However, unlike CTL, cytotoxicity was not MHC-restricted and the conventional T-cell receptor complex (CD3/Ti) appeared not to be involved in target cell recognition and cytolysis.     

Characterization of two sheep lymphocyte differentiation antigens, SBU-T1 and SBU-T6

The monoclonal antibodies 25.91 and 20.27 define two lymphocyte cell surface antigens of sheep. 25.91 is reactive with 60-80% of lymphocytes and 98% of thymocytes, and only stains surface immunoglobulin-negative peripheral lymphocytes. 25.91 immuno-precipitates a 67,000 MW protein from lymphocyte lysates under both reducing and non-reducing conditions, whereas immunoprecipitation of thymocyte lysates reveals a 67,000, 62,000 MW complex. The tissue distribution and molecular weight analysis reported here for the antigen recognized by 25.91 indicate that this antigen is the sheep homologue of the human T1 and mouse Ly 1 antigens. The monoclonal antibody 20.27 is reactive with 80% of thymocytes and the majority of cell surface immunoglobulin-positive peripheral blood lymphocytes (B cells), but is unreactive with peripheral blood T cells. 20-27 also stains Langerhans cells in skin tissue sections and large dendritic-like cells in the paracortex sections and large dendritic-like cells in the paracortex of lymph node tissue sections. Immunoperoxidase staining of thymus tissue sections with 20.27 shows intense staining of cortical thymocytes and an absence of staining within the medulla. Molecular weight analysis of the 20.27 antigen reveals two major bands of 46,000 and 12,000 MW under both reducing and non-reducing conditions. The 20.27 antigen has properties resembling MHC class I-like antigens such as T6 in the human and TL in the mouse.     

A monoclonal alloantibody detecting a polymorphism of the rat leucocyte common (LC) antigen

A monoclonal antibody, NDS 58, was produced by fusing the rat myeloma line Y3 Ag 1.2.3 with spleen cells from AS rats immunized with LEW thymocytes. The determinant recognized by this antibody is leucocyte-specific and present on all thymocytes, lymph node lymphocytes and some bone marrow cells. The strain distribution of NDS 58 was consistent with it being directed at the RT7a allele. Biochemical studies established that this antigen was carried on a cell surface glycoprotein with an apparent molecular weight on thymocytes of 190,000. This study confirms that the alloantigen originally designated Ly-1, and now termed RT7, is an allodeterminant of the LC molecule, and the controversy in this area, particularly with regard to the ART-1 alloantigen, is discussed.