Immunoperoxidase staining of surface and intracellular immunoglobulin in human neoplastic lymphoid cells.

An immunoperoxidase technique for the optical microscopic detection of cellular immunoglobulin has been used to stain fixed smears of human neoplastic B lymphoid cells. Only four out of 28 cases of chronic lymphatic leukaemia (CLL) showed membrane labelling by this technique. In contrast, when 14 samples from other types of B lymphoproliferative disorder (including hairy cell leukaemia, non-Hodgkin's lymphoma, and prolymphocytic leukaemia) were studied, all samples showed membrane immunoglobulin labelling (confirmed by capping experiments). This discrepancy was attributed to the greater density of surface immunoglobulin present on neoplastic cells in the latter group of disorders compared to CLL. This immunoperoxidase technique is therefore less sensitive than immunofluorescent staining of cells in suspension for the demonstration of neoplastic cell surface immunoglobulin. However, it offers a number of advantages (eg, excellent visualisation of cell morphology, permanence of stained preparations, and applicability to stored samples) which recommend it as the method of choice in certain clinical haematological contexts.     

Immunoglobulin determinants on the surface of lymphoid cells of carps.

Immunoglobulin determinants were detected by indirect immunofluorescence on the surface of numerous lymphocytes of perpheral blood, spleen, and pronephros of craps. The most interesting finding was the high proportion (65–68 %) of Ig⁺ lymphocytes in the thymus of early adult carps, possibly related to thymus function.     

HL-A antigens on man-mouse hybrid cells

Cultures of hybrid cells produced by fusion of human peripheral leukocytes with murine cell cultures were studied by means of mixed agglutination tests. All three hybrids studied contain human species-specific antigens. HL-A antigens corresponding to the phenotype of the leukocyte donor were demonstrated in two hybrids. With one of these hybrids, cloning experiments were performed. Five of six clones contained HL-A antigen controlled by one haplotype. In the remaining clone, which did not have human species-specific antigens, no HL-A antigens were detected. The third hybrid contained human alloantigens, but none of the HL-A antigens detected on the donor leukocytes.     

IgG2 and other immunoglobulin classes on the cell surface of rat lymphoid cells

The binding to rat lymphoid cells of the F(ab′)₂ fragments of purified rabbit antibodies specific for the Fc or Fd part of rat IgG₂ was studied. Both reagents heavily labeled about 15 % and 6 % of cells from spleen and cervical lymph nodes respectively of rats kept in a conventional animal house. In contrast to this there were very few IgG₂ positive cells in the thoracic duct lymph of any animals, or in the spleen of rats from an SPF animal house. Results on the quantitative binding of rabbit antibodies against rat Fab, IgM and IgA to rat lymphoid cells are also reported. IgM positive cells in the spleen bound much more anti-IgM relative to their binding of anti-Fab antibody, than IgM positive cells from thoracic duct lymph or cervical lymph node.     

Antibody-induced redistribution of HL-A antigens at the cell surface

The in vitro binding of anti-HL-A antibodies to the membrane of human lymphocytes induces important changes in the distribution of HL-A antigens on the cell surface. Following either direct or indirect immunofluorescence staining at 0 °C, cell-bound anti-HL-A antibodies are dispersed all over the cell surface. When the washed, stained lymphocytes are warmed and incubated at 37 °C, fluorescent antibodies cluster progressively at the cell surface. They form large spots of fluorescence, and sometimes single caps at one pole of the cell, outside which HL-A antigens are no longer detectable, but other antigens can still be found. Similar findings were made in electron microscopy, following indirect labeling of HL-A antigens on human lymphocytes, with ferritin or plant virus as the markers. With the indirect immunofluorescence technique, anti-human thymocyte antibodies and anti-mouse H-2 antibodies were found to induce a similar redistribution of the corresponding antigens. The mechanism and the interpretation of this displacement of surface antigens by antibodies are still unclear and are discussed in terms of membrane structure and immunological significance.     

The expression of immunoglobulin determinants on the surface of antigen-binding lymphoid cells in mice. II. Allotypes of the lg-1 locus

Inhibition of rosette-forming cells by anti-immunoglobulin sera has been used as a model system to investigate the phenotypic expression of IgG_2a allotypes on the surface of mouse spleen cells reactive against sheep erythrocyte antigens. The evidence presented suggests that reactive cells from mice heterozygous at the Ig-1 locus may not be restricted to the expression of the product of only one parental allele on their surface. Control studies with mixtures of cells from mice compatible with respect to H-2 antigens but differing at the Ig-1 locus suggest that this finding is not the result of a passive absorption phenomenon. This apparent lack of allellic exclusion was demonstrated with cells harvested during the early part of the response to a primary injection of antigen. Reactive cells later in the immune response appeared to be restricted to the expression of one parental allotype.     

Association of some cell surface antigens of lymphoid cells and cell surface differentiation antigens with early rat pregnancy

Monoclonal antibodies and the immunoperoxidase technique were used to localize some cell surface antigens of rat lymphoid cells and cell surface differentiation antigens on cryostat sections of early rat pregnancies. The W3/13 leucocyte sialoglycoprotein was detected almost constantly on trophoblast. The immunoglobulins were more associated with mother's rather than with embryo-derived tissues. We were unable to detect considerable amounts of class I and class II major histocompatibility complex-derived antigens on trophoblast and adjacent decidual cells. The Ia+ cells of the lymphocyte type were occasionally detected in the sites exhibiting presence of immunoglobulins. The Thy-1 cell surface differentiation antigen was detected on the cells producing Thy-1+ material among decidual cells. Depletion of Thy-1 was followed by the regression of decidualized tissue. The OX-2 antigen, known as minor glycoprotein of rat thymocytes, was detected on trophoblast cells and endothelia of decidual vessels, the latter exhibiting also class I major histocompatibility complex-derived antigens. The non-pregnant uterine tissues, as well as the oviduct epithelium were also investigated. The possible role of some of these antigens in the maintenance of the 'immunologically privileged' stage of trophoblast, and in the control of the rearrangement of maternal tissues surrounding the embryo, is discussed.     

Differential expression of HLA-DR and DR-linked determinants on human leukemias and lymphoid cells

Leukemic and normal hemopoietic cells were examined with the monoclonal anti-bodies DA2 and Genox 353 for the presence of HLA-DR and DR-linked (DC/MB) determinants, respectively. Although most non-T acute leukemias and leukemic cell lines expressed the monomorphic DR determinant detected by DA2, fewer than expected expressed the DR-linked polymorphic specificity detected by Genox 353. TdT+ lymphoid precursors from normal bone marrow were also DA2+ but Genox 353-. T cells and thymocytes which were DA2-, Genox 353- became DA2+, Genox 353+ after activation in vitro. Immunoprecipitation using DA2 and Genox 353 gave bands on polyacrylamide gel-electrophoresis which were of different molecular weights. In addition, DA2 could absorb out Genox 353 determinants from a cell lysate whereas Genox 353 could not absorb out DA2 determinants. It is concluded that DA2 and Genox 353 detect HLA-DR and DR-linked (DC1/MB1) determinants, respectively, and that these are differentially expressed on hemopoietic cells during differentiation.     

The expression of immunoglobulin determinants on the surface of antigen-binding lymphoid cells in mice. I. An analysis of light and heavy chain restrictions on individual cells

The expression of immunoglobulin light and heavy chain determinants on the surface of activated mouse spleen cells has been investigated using anti-immunoglobulin inhibition of antigen-binding reactions (rosette formation). The results indicate that the majority of binding cells appearing 4 to 7 days after a single injection of sheep erythrocytes have only one light chain type on their surface but simultaneously express more than one class of heavy chain class. As the response proceeds the proportion of heavy chain class “restricted” cells increases. Between days 15 to 30 following antigen injection the majority of binding cells have only one detectable heavy chain class on their surface. Control studies suggest that the appearance of more than one class of immunoglobulin on the surface of these cells is the result of active synthesis by the same individual cells and is not due to a passive absorption phenomenon. Experiments with antisera to non-immunoglobulin surface antigens indicate that the majority of antigen-binding cells inhibited by class specific sera are non, or minimally secreting B type lymphoid cells.     

The preparation and purification of HL-A monospecific antigens using platelets and various lymphoid sources

HL-A-active protein fragments have been isolated from lymphoid sources (tonsillar tissue, cells from lymphoblastoid cell lines (LCL cells)) and from platelets. The yields and specific activities of products obtained by enzymic chaotropic treatment indicate the superiority of 3 M KCl extraction of platelets. Individual HL-A antigenic specificities were further distinguished in fractions separated by polyacrylamide gel electrophoresis. Antigens were separated between the LA and FOUR loci, and within the LA locus. Purified monospecific HL-A antigenic preparations were conjugated with aminoethyl-Sepharose-4B, and used to remove corresponding specificities from alloantisera.     

Independence of chicken major histocompatibility antigens and tumor-associated antigen on the surface of herpesvirus-induced lymphoma cells.

Capping of chicken major histocompatibility (MHC) antigens on normal thymus, spleen, and peripheral blood leukocytes was demonstrated, although MHC antigens appeared to be present on only 15 to 18% of normal thymus cells. MHC antigen capping also occurred on cells from a Marek's disease herpesvirus-induced transplantable lymphoma (MDCT-NYM1). Capping of a Marek's disease tumor-associated surface antigen (MATSA) could be induced on MDCT-NYM1 lymphoma cells as well as on cells of two Marek's disease in vitro lymphoblastoid cell lines (MDCC-MSB1 and MDCC-LS1). Cocapping of MHC antigens and MATSA did not occur on MDCT-NYM1 lymphoma cells. The results suggest that MHC antigens and MATSA are not structurally associated on the cell membrane.