Kristof Liszka

VIL-A1, a monoclonal antibody reactive with common acute lymphatic leukemia cells.

The VIL-A1 monoclonal antibody raised against Reh cells reacts with common acute lymphatic leukemia (CALL) cells but not with normal or malignant B or T lymphocytes. It also shows no binding to normal or malignant myeloid, monocytic or erythroid cells, nor does it react with thrombocytes. The antibody is of IgM class and lyses CALL cells very efficiently in the presence of rabbit but not human complement. Immunoprecipitation experiments followed by SDS-polyacrylamide gel electrophoresis under reducing conditions revealed that VIL-A1 defines a 95,000 mol. wt membrane protein. Approximately 40% of it binds to lens culinaris lectin. Capping experiments showed that the membrane component defined by VIL-A1 co-caps with the one recognized by another recently described monoclonal antibody to CALL cells (J5).

Kinetics of activation antigen expression by in vitro-stimulated human T lymphocytes.

In this study a panel of monoclonal antibodies was used to investigate the kinetics of the appearance of activation-linked surface determinants as well as cytoplasmic and nuclear determinants in human T cells following lectin stimulation. Well known activation markers, such as Ia/DR, transferrin receptor, IL-2 receptor, T10, and gp24, were compared and investigated together with the T13 structure, recently found in this laboratory. T13, not demonstrable on resting T cells, could be seen within 24 hr after lectin stimulation. Kinetics of the appearance were similar to IL-2 receptor and transferrin receptor expression. Ia/DR synthesis was investigated separately for each polypeptide and the cytoplasmic invariant gamma-chain expression could be demonstrated for the first time with a gamma-chain-specific monoclonal antibody VIC-Y1. Moreover, gamma-chain synthesis seems to precede alpha- and beta-chain occurrence in human T cells. In addition, data from quantitative studies on antigenic densities are presented.

Monoclonal antibodies to human myelomonocyte differentiation antigens in the diagnosis of acute myeloid leukemia

The immunological definition of malignant lymphatic cells has already been routinely applied by many laboratories over a number of years. Today it is clear and undisputed that the phenotypic data obtained in that way are often very useful and supporting.The immunological definition of non-lymphoid leukemias is not yet as far advanced. Additional surface markers for the recognition of poorly differentiated non-lymphoid cells are clearly needed. In this paper ten monoclonal antibodies to myeloid surface antigens are described. It is hoped that they are a useful addition to the presently available relatively small panel of well studied myeloid surface markers which are displayed by immature malignant blast cells.

Typing of leukemic cells with monoclonal antibodies.

The immunofluorescence detection of surface-linked antigenic determinants by heteroantisera has greatly contributed to a better understanding of the heterogeneity of benign and malignant hemopoietic cells. The difficulties encountered in rendering these heteroantisera specific for a unique cell surface determinant have been a major drawback to a more rapid development of immunologic cell typing and, in particular, to a typing of non-lymphoid cells. The strategy ordinarily employed involved the extensive absorption of these heteroantisera with large numbers of cells of different cellular lineages. As a consequence, the resulting antibody titers were frequently low, thereby limiting the utility of these antisera for biological or clinical investigations. Finally, given the diversity of the immune response and the complexity of the absorption procedures, it has been difficult to reproduce antibodies with identical specificity, which frequently hindered the comparison of data obtained from different laboratories. With the introduction of the antibody-producing hybridoma technology by Kohler and M i l ~ t e i n , ~ most of these technical problems have been overcome. This technology provides for the first time uniform antibody reagents that are highly specific and of unlimited supply. The use of such antibodies should therefore considerably improve immunologic cell typing. Using this technology, we were able to raise a nkmber of monoclonal antibodies against cell type-specific differentiation antigens. These antibodies are of considerable help in the diagnosis and classification of human leukemia.

T14, A Non-modulating 150-Kd T Cell Surface Antigen

Monoclonal antibodies to human T lymphocyte surface structures have essentially contributed to a better understanding of cellular mechanisms and interactions involved in the generation of human T cell responses (1). Such antibodies have also proven useful in recent years in the diagnosis of autoimmune, immunodeficiency, and malignant diseases (2). Not surprisingly, monoclonal antibodies to human T lymphocytes are also increasingly used in the treatment of malignant diseases and for selective T cell elimination in bone marrow (3–5) and kidney transplantation (6–8).

Analysis of CD3-antibody-mediated inhibition of T-cell activation

In this study the influence of a non-mitogenic anti-CD3 antibody on accessory cell-dependent antigen and mitogen-induced T-lymphocyte proliferation has been investigated. The antibody was found to completely inhibit PHA, Con A, PWM, and tetanus toxoid stimulation, with no effect on the proliferation induced by the calcium ionophore A23187. VIT3 completely abrogated the production of IL-2 by lectin-stimulated T cells. It had no effect, however, on the IL-2-dependent proliferation of preactivated T-cell blasts. In addition, the antibody was able to elevate free cytoplasmic Ca2+ levels within minutes after the addition to T cells. Detailed time kinetic analyses revealed that the time interval critical for inhibition was significantly dependent on the interaction between T cells and accessory cells. Under standard conditions, in the presence of 10% non-T cells as accessory cells 50% inhibition was still achieved when VIT3 was added to PHA-stimulated T cells as late as 8 hr after the onset of culture. Delayed addition or a decrease in the number of added accessory cells significantly prolonged this time period. Lectin-stimulated T cells can thus obviously be inhibited via CD3 as long as they have not received all signals including those delivered by accessory cells. Although the underlying mechanisms are not clear so far, the observation that VIT3 at the same time triggers an early cytoplasmic Ca2+ response might indicate that it thereby actively interferes with antigen and lectin-initiated activation processes.

Leukemia Diagnosis with Monoclonal Antibodies

Summary With the introduction of hybridoma technology it became possible to obtain monoclonal antibodies and markedly improve immunologic cell-typing. We have, therefore, used this new technology for the production of monoclonal antibodies against human leukocyte-surface-antigens. This paper describes four cell-type specific monoclonal antibodies, which turned out to be very useful reagents in leukemia diagnosis. One of these antibodies, VIM-D5, is directed against a myeloid cell surface antigen. One, VIL-A1, is specific for the common acute leukemia associated antigen. VIB-C5 recognizes a B-cell differentiation antigen and VIE-G4 is specific for glycophorin-A and thus detects erythroid precursor cells.

Reactivity of Anti-Myeloid Monoclonal Antibodies with Committed Hematopoietic Precursor Cells

The development of monoclonal antibodies against surface structures of myeloid cells has contributed to the elucidation of differentiation processes at different levels of hematopoiesis. Most anti-myeloid antibodies have been defined for expression on cells at the stage of terminal myeloid maturation as well as on leukemic cell populations (for review, see Ref. 1). Only in relatively few studies, however, have antigenic structures of hematopoietic precursor cells been investigated with myeloid antibodies (2–8). In addition, the study of hematopoietic precursor cells has been so far mainly restricted to the granulo-monocytic cell lineage. Studies with clonal assays for erythropoietic and megakaryocytic stem cells are lacking in most publications. Furthermore, different in vitro culture techniques may complicate the interpretation of clonal assays even if the antibodies used are directed against the same antigenic structure. For example, a number of antibodies recognize the carbohydrate structure 3-fucosyl-N-acetyllactosamine [1G10 (4), R1B19 (5), VIM-D5 (9)] exposed on committed myeloid progenitor cells. Still the extent of lysis of CFU-GM subpopulations observed in different laboratories with one or the other of these antibodies varies considerably (4,5,8), the main reason probably being differences in the experimental design. For clonal assays of erythroid progenitor cells the situation is even worse since culture techniques using methylcellulose (10), plasma clot (11),or agar (12) are even more difficult to compare.