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M Samoszuk

Publications and source records attributed to M Samoszuk.

29 records · Page 2Linked to original sources
PubMed1991

Rapid detection of Epstein-Barr viral DNA by nonisotopic in situ hybridization. Correlation with the polymerase chain reaction.

A nonisotopic, in situ hybridization procedure is described for detecting Epstein-Barr viral (EBV) DNA in cytopreparations of pepsin-digested, unfixed cells in less than 3 hours. The procedure, which uses an EBV Bam HI W DNA probe conjugated directly to horseradish peroxidase, first was optimized using an EBV-infected cell line (ARH-77) and then validated on four positive control specimens and six negative control specimens. Optimization studies demonstrated distinct nuclear hybridization signal in less than 10% of formaldehyde-fixed, pepsin-digested ARH-77 cells. When formaldehyde fixation was omitted, more than 50% of the pepsin-digested ARH-77 cells had strong nuclear staining. Unfixed cells from the four positive controls had detectable hybridization signal; the presence of EBV-DNA in these specimens also was confirmed by a polymerase chain reaction. There was no hybridization signal in the six negative control specimens. The results indicate that this nonisotopic, in situ hybridization procedure rapidly detects the presence of EBV-DNA in cytopreparations of unfixed, pepsin-digested cells from clinical specimens. There is a tradeoff, however, between assay sensitivity and preservation of cytologic details.

B-Lymphocytes↗
PubMed1991

Frequent detection of Epstein-Barr viral deoxyribonucleic acid and absence of cytomegalovirus deoxyribonucleic acid in Hodgkin's disease and acquired immunodeficiency syndrome-related Hodgkin's disease.

Epstein-Barr viral DNA (EBV DNA) has been detected in 20 to 58% of Hodgkin's disease tumors analyzed by Southern blotting or polymerase chain reaction (PCR). Because patients with Hodgkin's disease are generally immunodepressed, it is possible that the EBV is not directly involved in the pathogenesis of Hodgkin's disease but is merely detectable by molecular techniques because of reactivation of a latent infection. The purpose of this study was to determine if EBV DNA could be detected in an even higher percentage of cases of Hodgkin's disease, including acquired immunodeficiency syndrome (AIDS)-related Hodgkin's disease, by using newly designed, PCR amplification primers, and to compare the incidence of EBV DNA with the incidence of another common, latent virus (cytomegalovirus) in Hodgkin's disease tissue. The PCR was performed on DNA extracted from cells from 15 benign hyperplastic lymph nodes and from 15 cryopreserved cases of Hodgkin's disease, including 2 cases of AIDS-related Hodgkin's disease. For negative controls, PCR was also performed without template DNA and on genomic DNA from E. coli, calf thymus, a murine myeloma, and from a human cell line. After 32 cycles of amplification, a 225 base-pair amplification product comigrating with an EBV-positive control was detected in none of the negative controls but was present in 14 out of 15 cases (93%) of Hodgkin's disease, including both cases of AIDS-related Hodgkin's disease, and in 2 out of 15 cases of benign lymphoid hyperplasia. By contrast, cytomegalovirus DNA was undetectable by PCR in any of our specimens. We conclude that in our study set, the PCR procedure detected EBV-DNA but not cytomegalovirus DNA in a high percentage of cases of Hodgkin's disease, including two cases of AIDS-related Hodgkin's disease. These findings strengthen the hypothesis that EBV may be involved in the pathogenesis of Hodgkin's disease and AIDS-related Hodgkin's disease.

Acquired Immunodeficiency Syndrome↗
PubMed1990

Detection of interleukin-5 messenger RNA in Reed-Sternberg cells of Hodgkin's disease with eosinophilia.

Hodgkin's disease of nodular sclerosis and mixed cellularity subtypes is frequently associated with eosinophilia. To determine if interleukin-5 (IL-5) is implicated in producing the eosinophilia, we performed in situ hybridization studies on cytopreparations of 16 cases of Hodgkin's disease with eosinophilia as well as cells from various controls. A single-stranded, anti-sense complementary DNA (cDNA) probe coding for a portion of the human IL-5 molecule was tail-labeled with digoxigenin -11-dUTP using terminal transferase, and then hybridized to messenger RNA (mRNA) within cells. An alkaline-phosphatase-conjugated antibody directed to digoxigenin was used with a chromogenic substrate to detect hybridized probe within cells. In all seven cases of nodular sclerosis subtype and nine cases of mixed cellularity subtype with eosinophilia, there was strong hybridization signal localizable to the cytoplasm of morphologically identifiable Reed-Sternberg cells and variants. Similar activity was detected only in rare cells from three normal spleens, and was undetectable in two cell lines used as a negative control and in one case of Hodgkin's disease without eosinophilia. Pretreatment of the cytopreparations with the RNase inhibitor diethylpyrocarbonate greatly increased the hybridization signal. Based on this controlled study, we conclude that mRNA coding for IL-5 is expressed in Reed-Sternberg cells and variants. This observation may explain the eosinophilia associated with Hodgkin's disease and provide insight into the origin of the Reed-Sternberg cell.

Eosinophilia↗
PubMed1989

Shared idiotypes expressed by human B-cell lymphomas.

Each B-cell lymphoma expresses a surface immunoglobulin that contains unique antigenic determinants (idiotypes). We have produced 199 murine monoclonal antibodies reactive with the idiotypes isolated from 67 patients with follicular small-cleaved-cell lymphoma. These antiidiotype antibodies were analyzed for their ability to react with lymphoma cells from patients other than the one against which each antibody was made. Twenty of the 199 antiidiotype antibodies were reactive with lymphoma cells from more than one patient. Depending on the antibody, the frequency of idiotype sharing ranged from 0.6 to 6.2 percent of B-cell lymphoma tumors evaluated. Tumors could be grouped into distinct families on the basis of their reactivity with these antibodies. In the aggregate, the 20 antibodies reacted with a total of 49 of 150 B-cell lymphomas (33 percent), including 30 of 110 follicular small-cleaved-cell lymphomas (27 percent). Many of these shared idiotypes were expressed by more than one histopathological subtype of lymphoma. We conclude that a panel of antibodies reactive with shared idiotypes can be produced for patients with B-cell lymphoma, obviating the need to produce an antiidiotype antibody for each patient.

Antibodies, Anti-Idiotypic↗
PubMed1987

Screening for anti-idiotypic monoclonal antibodies on paraformaldehyde-fixed lymphoma cells.

Paraformaldehyde-fixed, human lymphoma cells on glass slides were used to screen murine hybridoma supernatants for anti-idiotypic monoclonal antibodies by a rapid indirect immunofluorescence technique. The method is shown to require far fewer cells for screening than current techniques, and it provides results that correlate with the results of screening by flow cytometry or by capture ELISA. We conclude that paraformaldehyde fixation preserves the idiotypic determinants of lymphomas. The assay described in this report, therefore, has significant advantages over current methods for producing anti-idiotypic monoclonal antibodies.

Animals↗
PubMed1987

Immunohistochemical characterization of a 183 KD myeloid-specific-DNA-binding protein in B5 fixed, paraffin-embedded tissues, and bone marrow aspirates by monoclonal antibody BM-1.

A monoclonal antibody, designated BM-1, which is reactive in B5 formalin-fixed, paraffin-embedded tissues, has been generated against a cytoplasmic and nuclear antigen expressed in human myeloid precursor cells and derived leukemias. Using the avidin-biotin-complex immunoperoxidase procedure, BM-1 was found to stain selectively myeloid precursor cells in normal bone marrow and mature granulocytes in the blood. In a screen of 26 normal adult and fetal human organs fixed in B5 formalin, BM-1 was negative in all nonhematopoietic tissues with the exception of tissue granulocytes and scattered cells in the peripheral cortex of the thymus. Likewise a screen of 30 solid tumor cell lines including a spectrum of carcinomas, sarcomas, and neural-derived tumors was negative. BM-1 was also negative with 21 T and B cell lymphomas and 11 Hodgkin's disease tumors. A preliminary study of tumors of the hematopoietic system revealed that BM-1 was reactive with M2 and M3 acute myelogenous leukemias (AML), chronic myelogenous leukemias (CML) and myelomonocytic leukemias, and granulocytic sarcomas. M1, M4, M5, and M6 AML clot preparations were negative in this study, indicating that BM-1 may have a role in the histopathologic diagnosis of myelogenous leukemia. Myeloid leukemic cell lines HL-60, ML-2, KG1, and TPH-1-O showed BM-1 nuclear and/or cytoplasmic reactivity in a subpopulation of cells, but erythroid and lymphoid leukemias and all lymphoma cell lines were negative. Immunoperoxidase studies of a panel of fetal tissues showed BM-1 positive cells in the peripheral cortex of the thymus and portal myelopoietic regions of the liver at 18 weeks gestation. Finally, DNA-cellulose and solid phase radioimmunoassay (RIA) techniques developed in our laboratory demonstrate that the BM-1 antigenic domain is reactive only after binding to eukaryotic but not prokaryotic single- or double-stranded DNA. Immunoblot techniques using a DNA-cellulose purified protein sample revealed that BM-1 recognizes a 183 kD protein. These studies indicate that BM-1 is recognizing a myeloid-specific antigen that, because of its DNA binding characteristics, may have an important role in the differentiation of myeloid cells at the molecular level.

Animals↗
PubMed1987

Eosinophil peroxidase is detectable with a monoclonal antibody in collagen bands of nodular sclerosis Hodgkin's disease.

An IgG2a murine monoclonal antibody derived against human eosinophils was shown to immunoprecipitate the 78,000 dalton form of human eosinophil peroxidase (EPO). To confirm the specificity of the antibody, we used a glucose-oxidase avidin biotin procedure to immunostain 32 human cell lines and tissues. An eosinophilic subline of HL-60 promyelocytic leukemia was the only cell type other than eosinophils to be recognized by the antibody. Because previous reports have described occult eosinophilic degranulation in tissues with a variety of pathological conditions, we immunostained cryostat sections of four consecutive lymph node biopsies of nodular sclerosis Hodgkin's disease with the monoclonal antibody. Our objective was to characterize by immunohistology the eosinophilic infiltration in a lymphoma that frequently contains substantial numbers of eosinophils. In all four cases of nodular sclerosis Hodgkin's disease, there was striking and extensive deposition of EPO in a dendritic pattern throughout the connective tissue and collagen bands. The extent of deposition of EPO in the bands far exceeded the degree of infiltration by intact eosinophils, as determined by examination of routinely stained tissue sections. A similar dendritic pattern was not observed in any of six benign lymph nodes that were immunostained for EPO. We conclude that the monoclonal antibody described in this report is specific for EPO and that eosinophils extensively degranulate and release EPO in the bands of nodular sclerosis Hodgkin's disease. Moreover, the degree of eosinophilic infiltration in this disorder cannot be assessed solely on the basis of intact eosinophils.

Antibodies, Monoclonal↗
PubMed1983

Serum theophylline assay by rate nephelometric inhibition immunoassay: a comparison with high-performance liquid chromatography.

The reliability of two clinical laboratory methods for assaying theophylline in serum is compared. Analysis of serum-based standards and 70 patient specimens by rate nephelometric inhibition immunoassay (NIIA) and by high-performance liquid chromatography (HPLC) demonstrates that both methods have comparable random analytical variation. The results indicate that NIIA is a reliable method for measuring serum theophylline and may serve as a practical alternative to the HPLC method.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

In vitro and in vivo interactions of D-penicillamine with tumors.

D-penicillamine (PSH) is a copper chelator that generates hydrogen peroxide and inhibits neovascularization. As hydrogen peroxide is toxic to some tumor cells and to blood vessels, we reasoned that PSH plus copper would inhibit tumors in vivo and in vitro. To test this hypothesis, we first incubated murine J558L plasmacytoma cells with varying combinations of drug (PSH and/or copper sulfate) plus modulators (fetal calf serum, dithiothrietol, catalase, eosinophil peroxidase) and then used fluorescence microscopy to measure cell proliferation, necrosis, and apoptosis. We also incubated various types of human tumor cells with PSH plus copper for 24 hours and then measured the number of surviving cells 24 hours later. For the in vivo studies, we measured the effects of 7 daily i.p. injections of 10 mg of PSH on the growth rates of interleukin-5 genetransfected J558L tumors in 20 BALB/c mice. Our experiments demonstrated that PSH plus copper exerted a significant antiproliferative effect on tumor cells in vitro that was neutralized by protein or catalase and enhanced by adherent eosinophil peroxidase. Human acute myelogenous leukemia cells were especially sensitive to PSH plus copper. In vivo, however, PSH had no significant effect on the growth rates of J558L tumors that were infiltrated by eosinophils. We conclude that the interaction of PSH-copper with tumors is primarily antiproliferative, mediated by hydrogen peroxide, and inhibitable by protein. Therefore, for PSH to be an effective antineoplastic drug strategies will need to be developed to prevent its rapid neutralization by protein.

Animals↗