Granulocyte-specific Monoclonal Antibody Research Articles

Neutrophils prevent extracellular colonization of the liver microvasculature by Salmonella typhimurium.

The early course of hepatic infection with the facultative intracellular bacterial pathogen Salmonella typhimurium was examined in control mice and in mice selectively depleted of neutrophils by treatment with a granulocyte-specific monoclonal antibody. The results show that >200-fold more salmonellae were recovered in livers of the latter group of mice than in livers of the former group by 24 h of parenterally initiated infection. Comparative histological examination of the livers from both groups of mice indicated that neutrophils participate in early anti-Salmonella defense in the liver in part by aborting infection in permissive hepatocytes and by inhibiting extracellular bacterial colonization of the hepatic microvasculature. It is shown in addition that systemic salmonellosis was also severely exacerbated in neutropenic mice infected intragastrically with the pathogen.

Neutrophils are critical for host defense against primary infection with the facultative intracellular bacterium Francisella tularensis in mice and participate in defense against reinfection.

It is generally believed that immunity to experimental infection with the facultative intracellular bacterium Francisella tularensis is an example of T-cell-mediated immunity that is expressed by activated macrophages and mediated by Francisella-specific T cells. According to the results presented herein, neutrophils are also essential for defense against primary infection with this organism. It is shown that mice depleted of neutrophils by treatment with the granulocyte-specific monoclonal antibody RB6-8C5 are rendered defenseless against otherwise sublethal doses of F. tularensis LVS inoculated intravenously or intradermally. In neutrophil-depleted mice, the organism grew progressively in the livers, spleens, and lungs to reach lethal numbers, whereas infection was resolved in normal mice. Although neutrophils were found to resistance to reinfection, their participation was less important. The results suggest that neutrophils are needed for defense against primary infection because they serve to restrict the growth of F. tularensis before it reaches numbers capable of overwhelming a developing specific immune response. The exact way that neutrophils achieve this is not clear at this time, although it is probable that they contribute in ways other than by ingesting and killing the bacterium.

Granulocyte-specific monoclonal antibody technetium-99m-BW 250/183 and indium-111 oxine-labelled leucocyte scintigraphy in inflammatory bowel disease

Granulocyte-specific monoclonal antibody technetium-99m-BW 250/183 and indium-111 oxine-labelled leucocyte scintigraphy in inflammatory bowel disease

Granulocyte-specific monoclonal antibody technetium-99m-BW 250/183 and indium-111 oxine-labelled leucocyte scintigraphy in inflammatory bowel disease.

Thirty-three patients suspected of suffering from inflammatory bowel disease were studied. Autologous leucocytes were labelled with indium 111 oxine and re-injected simultaneously with 0.3-0.5 mg of technetium 99m granulocyte-specific monoclonal antibody BW 250/183. Two scans were obtained, the early scan 3-4 h postinjection (p.i.) and the late scan 18-24 h p.i. Using the endoscopy study as standard, the diagnostic accuracy of both agents was determined. Sensitivity, specificity and accuracy of 111In scans was 88.8%, 100.0% and 93.7% at 4 h and 94.7%, 100.0% and 96.9% at 24 h, respectively. Concerning the results using antibodies, the values were 61.1%, 100.0% and 78.1% at 4 h and 78.9%, 92.8% and 84.8% at 24 h, respectively. Segmental analysis showed concordance in 89.3% and 93.3% of the cases at 4 and 24 h, respectively. Though less sensitive and less accurate than scanning employing indium 111 leucocytes, BW 250/183 granulocyte-specific scintigraphy can be used for inflammatory bowel disease diagnosis and localization.

Cytohistologic and immunohistochemical findings in Hodgkin's disease, mixed cellularity type, with a high content of epithelioid cells.

We studied 123 lymph node biopsies from 99 patients with Hodgkin's disease, mixed cellularity type, with a high content of epithelioid cells (HDMCep), by light microscopy using conventional histologic and immunohistochemical techniques. The cellular composition and histologic structure, which are described in detail, serve as a basis for discriminating HDMCep from similar lymphomas with a high content of epithelioid cells, especially from lymphoepithelioid cell lymphoma (Lennert's lymphoma, LeL) and angioimmunoblastic (LgX) type of peripheral T-cell lymphoma with a high content of epithelioid cells (AILD-PTCLep). In all lymph nodes from patients with HDMCep, the nodal architecture was effaced. In 11% it was partially replaced by a massive infiltration of epithelioid cells and small to medium-sized lymphoid cells occurring in varying proportions with some immunoblasts. In all biopsy specimens, some typical Sternberg-Reed (SR) and Hodgkin (H) cells were found. Plasma cells and plasma cell precursors were present in all biopsy specimens; in most cases, they were sparse to moderate in number. Eosinophils were always present; half of the biopsy specimens had rather large numbers of them. Formation of granulomas like those seen in sarcoidosis was noted in 7% of the specimens. Increased vascularity and an increased number of fibers were found in 53% and 63% of the biopsy specimens, respectively. Necrosis was seen in 20% of the specimens. Immunohistochemically in 30 of 67 (45%) biopsy specimens, giant cells stained positively for the granulocyte-specific monoclonal antibody 3C4 (approximately CD15). Plasma cells and plasma cell precursors in all 70 specimens exhibited a polyclonal Ig pattern. A comparison of the main clinical and laboratory data in these three entities reveals both similarities and differences. HDMCep--defined as a special variant of Hodgkin's disease, mixed cellularity type (HDMC)--marks apparently the border between Hodgkin's disease and non-Hodgkin's lymphomas.

Quantitation of conjugate formation between human polymorphonuclear leukocytes and antibody-coated target cells by flow cytometry: the role of Fc receptor and LFA-1 antigen.

The specific binding of human polymorphonuclear leukocytes (PMN) to antibody-coated target cells was characterized by flow cytometry. PMN were labeled with phycoerythrin-E (PE) via a granulocyte-specific monoclonal antibody (leu-M1) and mixed with fluorescein isothiocyanate-labeled K562 tumor cells sensitized with rabbit antiserum. Specific conjugates were formed as analyzed by two-color fluorescence in a flow cytometer. The formation of stable conjugates was dependent on initiation of contact, temperature, time, and antiserum concentration. Studies with inhibitors implicate that microfilaments, but not microtubules, Ca2+, Mg2+, or energy-dependent processes were a prerequisite for binding of PMN to the antibody-coated target cells. No conjugates were formed when uncoated target cells were used or when the experiment was performed in the presence of protein A, indicating that binding was specifically mediated through Fc receptors (FcR). Monoclonal antibodies against the FcRII and FcRIII were used to address the role of these receptors in conjugation. One of the two anti-FcRIII antibodies and an anti-FcRII antibody effectively prevented conjugation. A monoclonal antibody directed against the common beta-chain of the adhesion molecule family and a combination of antibodies against the alpha-chain of LFA-1 and Mo-1 also blocked conjugation when target cells were sensitized under suboptimal conditions. The antibody against the beta-chain also diminished killing of antibody-coated K562, as measured by chromium release when included in the cytotoxicity assay. These results indicate that flow cytometry permits accurate quantitation and characterization of the binding between PMN and antibody-coated target cells, which in principle, can be prevented by monoclonal antibodies against surface receptors. Binding is primarily established by both the FcRII and FcRIII. Adhesion-associated molecules on the PMN surface contribute to optimal binding.

Histological and Immunohistological Findings in Lymphoepithelioid Cell Lymphoma (Lennertʼs Lymphoma)

Four hundred and two cases considered or suspected to be lymphoepithelioid cell lymphoma (Lennert's lymphoma) were analyzed morphologically, immunohistochemically, and clinically. One hundred and eight of these cases, investigated in 180 biopsies, fulfilled the morphological and immunohistochemical criteria for lymphoepithelioid cell lymphoma and are herein reported. Cellular composition and histological structure are described in detail as a basis for discriminating Lennert's lymphoma from similar lymphomas with a high content of epithelioid cells (Hodgkin's disease, AILD type of T-cell lymphoma, lymphoplasmacytic lymphoma) and from inflammatory epithelioid cell reactions. Single typical Hodgkin cells were found in only 3.8% of biopsy specimens examined, and single typical Sternberg-Reed cells were found in only 2.2% of the biopsy specimens examined. Because these single Hodgkin and Sternberg-Reed cells were situated in a relatively monotonous lymphoid cellular pattern and because both these cells types also may occur in peripheral T-cell lymphomas, we include cases with such cells in the category of Lennert's lymphoma. Eight percent of the patients with lymphoepithelioid cell lymphoma showed transformation into a large-cell T-cell lymphoma without the prominent focal epithelioid cell component previously observed. Immunohistochemically, seven of the 69 biopsy specimens with detectable giant cells stained positively for the granulocyte-specific monoclonal antibody 3C4 (approximately CD15). Plasma cells were rare and always showed a polytypic immunoglobulin pattern. Lymphoepithelioid cell lymphoma, defined as a lymphoma of CD4+ T lymphocytes, marks the border between Hodgkin's disease and the non-Hodgkin's lymphomas. Today, absolute criteria for distinguishing between these two classes of lymphoma are lacking.

Granulocyte-specific monoclonal antibody inhibiting cytotoxicity reactions in the chicken

Monoclonal antibody (MAb) 10, specific for chicken granulocytes, is described for the first time. Treatment of peripheral blood leukocytes with this MAb markedly decreased natural cytotoxicity reaction against the target cell line. The antibody-dependent cellular cytotoxicity (ADCC) activity of purified granulocytes was also severely affected by treatment with MAb 1 C3. These results suggest that 1 C3 detects a functional surface antigen on chicken granulocytes and support the hypothesis that granulocytes are the main effector cells in natural cytotoxicity in the chicken.

The role of interferon in the adenovirus-induced augmentation of granulocyte-mediated cytotoxicity in chicken

The effects of human adenoviruses on the granulocyte-mediated natural cytotoxicity of chicken leukocytes were investigated. A significant, but transient augmentation of granulocyte cytotoxicity was observed 24 h after virus injection, followed by a relatively long period of its suppression. A good correlation was found between the augmented cytotoxicity and interferon induction. The interferon-inducing capacity of adenovirus type 6 and type 12 in vitro similarly ran parallel with their ability to stimulate granulocyte-mediated Cytotoxicity. An adenovirus-induced elevation of cytotoxicity was not observed when IFN production was inhibited by pretreatment of the leukocytes with monoclonal antibody specific for bursal cells and monocytes. In addition, anti-IFN antibody abrogated the stimulation of cytotoxicity as well. During the in vitro experiments in which granulocyte-specific monoclonal antibody was applied, evidence was found that the effector cell activity is associated with the granulocytes. These results suggest that both the in vitro and the in vivo adenovirus-induced augmentation of granulocyte-mediated cytotoxicity is due to the IFN-inducing capacity of the virus. In chickens, the rapid augmentation of the granulocyte cytotoxicity may be important in the acute stage of infection, increasing the resistance to the virus in question and also to bacterial infections.

Characterization of cell surface glycoproteins recognized by the granulocyte-specific monoclonal antibody, AHN-1

Major surface-iodinated proteins of M r 105,000 and 145,000 of normal human neutrophils are immunoprecipitated by a number of monoclonal antibodies (AHN-1 to AHN-6), which react specifically with granulocytes among peripheral blood cells and selectively inhibit phagocytosis. These proteins, and an M r 60,000 component, were purified by monoclonal antibody affinity chromatography, molecular sieve chromatography, and preparative polyacrylamide gel electrophoresis. Each of the three purified proteins was immunoprecipitated by all six antibodies. Nevertheless, tryptic peptide maps of the three proteins indicated that each was a distinct component. AHN-1 to AHN-6 also bound to glycolipid fractions of human neutrophils, and the binding of each antibody to human neutrophils was blocked by the carbohydrate sequences, lacto- N-fucopentaose III. The data indicate that a predominant antigenic determinant of human neutrophils is lacto- N-fucopentaose III, or related carbohydrates, present on three distinct proteins as well as glycolipids. At least one of these molecules appears to be involved in the process of phagocytosis.

Monoclonal Antibody AHN-1 Inhibits Phagocytosis by Human Neutrophils

The granulocyte-specific monoclonal antibody, AHN-1, immunoprecipitates two major surface-iodinated proteins of 105,000 and 145,000 to 150,000 daltons from normal human neutrophils. In this study, the effect of AHN-1 on a number of neutrophil functions was evaluated in vitro. Both complement- and antibody-mediated phagocytosis were inhibited when human neutrophils were pretreated with AHN-1 and opsonized bacteria were used as targets. The inhibition of phagocytosis was specific, in that lysosomal enzyme release and chemotaxis were not altered by treatment with AHN-1. AHN-1 did inhibit superoxide production by neutrophils in response to particulate stimuli, but not in response to the soluble stimulus, 12-O-tetradecanoylphorbol-13-acetate. The data indicate that one or both of these surface proteins may be important in the process of phagocytosis. AHN-1 should be useful in isolating and further characterizing the nature of these molecules.

Monoclonal antibody AHN-1 inhibits phagocytosis by human neutrophils

The granulocyte-specific monoclonal antibody, AHN-1, immunoprecipitates two major surface-iodinated proteins of 105,000 and 145,000 to 150,000 daltons from normal human neutrophils. In this study, the effect of AHN- 1 on a number of neutrophil functions was evaluated in vitro. Both complement- and antibody-mediated phagocytosis were inhibited when human neutrophils were pretreated with AHN-1 and opsonized bacteria were used as targets. The inhibition of phagocytosis was specific, in that lysosomal enzyme release and chemotaxis were not altered by treatment with AHN-1. AHN-1 did inhibit superoxide production by neutrophils in response to particulate stimuli, but not in response to the soluble stimulus, 12-O-tetradecanoylphorbol-13-acetate. The data indicate that one or both of these surface proteins may be important in the process of phagocytosis. AHN-1 should be useful in isolating and further characterizing the nature of these molecules.