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Biomedical subjects

M A Epstein

Publications and source records attributed to M A Epstein.

At least 73 records · Page 4Linked to original sources
PubMed1984

A highly sensitive enzyme-linked immunosorbent assay to quantitate antibodies to Epstein-Barr virus membrane antigen gp340.

An enzyme-linked immunosorbent assay (ELISA) has been developed for the detection of antibodies to Epstein-Barr (EB) virus membrane antigen (MA) glycoprotein, gp340, in tamarins. The assay was found to be a thousand-fold more sensitive than conventional indirect immunofluorescence tests and consequently it was possible to follow accurately the sequential production of specific antibodies to gp340 by tamarins during a course of immunization.

Animals↗
PubMed1984

A structural investigation of the Epstein-Barr (EB) virus membrane antigen glycoprotein, gp340.

Epstein-Barr (EB) virus membrane antigen (MA) glycoprotein (gp340) purified by a molecular weight-based technique has been subjected to biochemical analysis. Following treatment with glycosidases or tunicamycin during synthesis, the carbohydrate moiety was found to be made up of both O-linked and N-linked types and to constitute about 50% of the molecular mass. Digestion studies with neuraminidase and oligosaccharidase have indicated that the molecule is heavily sialated with most of the sialic acid located on the O-linked sugars. The high carbohydrate content of gp340 appears to confer resistance to proteolysis; thus, V8 protease was only effective at concentrations above 1 mg/ml when three large fragments of mol. wt. 330K, 190K and 160K were generated. Removal of sialic acid before V8 protease digestion did not alter this pattern nor affect the antigenicity of the digestion fragments. Antigenicity of the intact molecule was likewise unaffected by removal of sialic acid nor were the O-linked and N-linked carbohydrate moieties essential for this property. The binding of virus-neutralizing human sera and monoclonal antibody by gp340 from which either O-linked or N-linked sugars had been removed seems to indicate that the sites on the molecule that generate the neutralizing antibodies are present in the protein component. The significance of these results is discussed in relation to the development of a subunit vaccine against EB virus.

Animals↗
PubMed1983

Cross-reactivity of self-HLA-restricted Epstein-Barr virus-specific cytotoxic T lymphocytes for allo-HLA determinants.

Epstein-Barr (EB) virus-specific cytotoxic T cells, prepared from virus-immune donors by reactivation in vitro and maintained thereafter as IL-2-dependent T cell lines, have been tested against large panels of EB virus-transformed lymphoblastoid cell lines of known HLA type. Whilst the pattern of lysis of the majority of targets was always consistent with HLA-A and HLA-B antigen restriction of effector function, in several cases it was noticed that certain HLA-mismatched targets were also reproducibly lysed. When this "anomalous" lysis was investigated in detail, it was found to be directed against allodeterminants on class I HLA antigens; thus, mitogen-stimulated as well as EB virus-transformed lymphoblasts from the relevant target cell donors were sensitive to the killing, and in each case the lysis could be specifically blocked by monoclonal antibodies to class I HLA antigens. In one example the target for this alloreactive lysis could be identified as a single serologically defined antigen, HLA-Bw57, while in another example lysis was directed against a "public" epitope common to HLA-Bw35, -Bw62, and a subset of -B12 antigens. Both cold target inhibition experiments and limiting dilution analysis strongly suggested that this alloreactive lysis was being mediated by the same effector T cells that recognize EB viral antigens in the context of self-HLA. This is the first demonstration in man that alloreactive responses can be derived from within the antigen-specific, self MHC-restricted T cell repertoire.

Antibodies, Monoclonal↗
PubMed1983

Epstein-Barr virus-specific cytotoxic T lymphocytes as probes of HLA polymorphism. Heterogeneity of T cell-restricting determinants associated with the serologically defined HLA-A2 antigen.

Epstein-Barr (EB) virus-specific effector T cell lines were established from nine virus-immune donors positive for the serologically defined HLA-A2 antigen; of these, four lines contained a demonstrable A2-restricted cytotoxic component. When these four effector populations were each tested on the same panel of EB virus-transformed lines from 20 HLA-A2-positive individuals, 16 of the target cell lines were consistently killed at levels above 25% of the relevant autologous cell lysis. Cytotoxicity appeared to be mediated through a restricting determinant associated with the 'common A2' antigen that these lines shared; indeed the lysis could be specifically blocked by high concentrations of an HLA-A2-specific monoclonal antibody. In contrast, 4 out of 20 target cell lines were not killed by HLA-A2-restricted effector cells, even though they did express the serologically defined A2 antigen and were found in other tests to be susceptible to EB virus-specific cytolysis restricted through other HLA-A or -B antigens on their surface. These results suggest that EB virus-specific cytotoxic T cells can distinguish between serologically identical HLA-A2 molecules via the heterogeneity of their T cell-restricting determinants. Data from one of the effector cell populations further suggested that a serologically defined cross-reaction between the otherwise distinct HLA-A2 and -Bw57 antigens might also be reflected in a cross-reactivity of T cell-restricting determinants.

Adult↗
PubMed1983

A culture method giving substantial yields of normal nasopharyngeal epithelial cells for work with Epstein-Barr virus.

A culture method, utilising a feeder layer of lethally irradiated 3T3 fibroblasts and medium supplemented with hydrocortisone, cholera toxin, and epidermal growth factor, has been elaborated for the in vitro growth of normal human nasopharyngeal epithelial cells. This method allowed the cells to be grown in vitro for periods of up to 146 days, very considerably longer than in previously reported studies, and ensured that the cultures remained largely free from contaminating human fibroblasts. It was found possible to subculture the nasopharyngeal epithelial cells through numerous passages both by dispersing monolayers into single cell suspensions and by transferring coverslip monolayers of the cells to individual Petri dishes. By combining these two methods, at least 50 replicate epithelial cultures could be produced from each tissue sample, thus providing for the first time cultured nasopharyngeal epithelial cells in quantities suitable for extensive experiments with Epstein-Barr virus.

Cells, Cultured↗
PubMed1983

Selective reactivation of Epstein-Barr virus-specific cytotoxic T cells by stimulation in vitro with allogeneic virus-transformed HLA-homozygous typing cells.

Epstein-Barr (EB) virus-specific cytotoxic T-cell preparations, produced by stimulation in vitro of peripheral blood lymphocytes with the autologous virus-transformed cell line, are HLA-A and B antigen-restricted and, with some donors, show preferential restriction through one or two of the four relevant antigens of the donor's HLA type. It has now been demonstrated that such EB virus-specific cytotoxic T cells may also be reactivated by stimulation with allogeneic virus-transformed cells provided that there is no mismatch of the HLA-A and B antigens between the responder and stimulator cell donors. In particular, virus-transformed cell lines from HLA-homozygous donors HLA-A and B antigen-matched to one of the haplotypes of an HLA-heterozygous responder were shown to reactivate selectively only those EB virus-specific cytotoxic T cells restricted through the HLA-A and B antigens present on the allogeneic stimulating cells. In addition to confirming the polyclonal nature of the HLA-restricted EB virus-specific cytotoxic T-cell response, this new experimental procedure has allowed the production, and subsequent expansion as cell lines dependent upon T-cell growth factor, of those effector cells restricted through the "nonpreferred" HLA antigens that are poorly represented in the response induced by stimulation with autologous virus-transformed cells.

Cell Line↗
PubMed1983

Purification and properties of the gp340 component of Epstein-Barr virus membrane antigen in an immunogenic form.

Preparative SDS-polyacrylamide gel electrophoresis has been used in the purification of the gp340 component of Epstein-Barr (EB) virus-determined membrane antigen (MA), in tractable quantities, from the B95-8 marmoset lymphoblastoid cell line. Successful renaturation of the purified molecule was achieved. This procedure gave a 50-fold increase in the recovery of antigen compared to conventional techniques. The data suggest that the antigenic sites recognized by human sera containing antibodies to MA are largely confined to the protein portion of the molecule. An eightfold improvement in the yield of gp340 was obtained when B95-8 cells were cultured in the presence of 12-O-tetradecanoyl-phorbol-13-acetate. Gel filtration studies indicate that the major polypeptide components of MA are not associated in detergent solution. Immunization of rabbits with purified and renatured gp340 resulted in the generation of high-titre antisera which were specific for gp340, demonstrating that antigen prepared by this procedure is suitable for further evaluation as an experimental vaccine against EB virus infection.

Animals↗
PubMed1983

Factors influencing population growth of a colony of cotton-top tamarins.

43% of full-term births in a colony of Saguinus oedipus oedipus occurred during April and May. Interbirth interval was usually 12 months in females which reared live young and 7 months in those whose young died perinatally. 81% of all full-term births were of twins, the rest were singles. High mortality among neonates was seen with a group of bought-in animals which had been taken from their parents at an early age without participating in the rearing of siblings, and was attributable to parental incompetence. Colony management aiming to ensure that animals gain experience in caring for young before breeding is described. Mortality among all animals older than 7 days was very low. Marginal protein deficiency may have been involved in a chronic diarrhoea and weight loss syndrome seen in some animals. They recovered after addition of hard-boiled egg to the diet.

Animal Husbandry↗
PubMed1982

Interaction of Epstein-Barr virus with leukaemic B cells in vitro. II. Cell line establishment from prolymphocytic leukaemia and from Waldenström's macroglobulinaemia.

Leukaemic B-cell populations were prepared from six patients with high-count prolymphocytic leukaemia (PLL) as well as from one patient with Waldenström's macroglobulinaemia (WM) in frankly leukaemic phase, and their response to in vitro Epstein-Barr (EB) virus infection was monitored in terms of expression of the virus-associated nuclear antigen EBNA and of virus-induced transformation to continuous cell lines. The individual leukaemic populations, tested on several occasions, gave reproducibly different responses one from another which were not obviously related to differences either of surface immunoglobulin phenotype or of immunoglobulin secretor status in vivo. After infection, four out of six PLL populations showed either transient or a more persistent expression of EBNA, always involving a minority of the cells, with no evidence of any virus-induced transformation up to six weeks. In contrast, two out of six PLL samples as well as the WM sample rapidly gave rise to EBNA-positive cell lines which, on the evidence both of restricted immunoglobulin class expression and of abnormal marker chromosomes, were clearly derived from the leukaemic cells. Further comparative studies of such leukaemic B-cell populations may help to define host cell components necessary for the triggering of EB-virus-induced cellular transformation.

Animals↗
PubMed1982

Epstein-Barr-virus-specific T-cell memory in renal-allograft recipients under long-term immunosuppression.

The level of long-term responsiveness of cytotoxic T cells to Epstein-Barr (EB) virus has been measured in peripheral-blood mononuclear-cell cultures from symptom-free renal-allograft recipients under conventional immunosuppression. 8 of 22 patients showed a profoundly impaired responsiveness when compared with health control donors. The allograft recipients as a group showed raised antibody titres to EB-virus capsid and early antigens, but within the group there was no clear relation between these antibody titres and the level of EB-virus-specific cytotoxic-T-cell function. These findings are particularly relevant to the pathogenesis of the malignant lymphomas to which renal-allograft recipients are unusually susceptible, in view of the evidence implicating EB virus.

Adult↗
PubMed1982

Monoclonal antibodies to Epstein-Barr virus-induced, transformation-associated cell surface antigens: binding patterns and effect upon virus-specific T-cell cytotoxicity.

Spleen cells from mice immunized with Epstein-Barr virus-transformed lymphoblastoid cells (EB-LCL) were used to generate monoclonal antibodies to cell surface antigens associated with the EB virus-transformed state. Radioimmune and immunofluorescence binding assays identified two antibodies, MHM6 and AC2, which reacted consistently with all EB-LCL tested, with a subpopulation of cells in some but not all EB virus genome-positive Burkitt lymphoma lines, but with none of a range of EB virus genome-negative cell lines of lymphoma or leukaemia origin. While MHM6 appeared to bind an EB virus-related antigen, AC2 bound some other cell surface antigen which was also found on a small subpopulation of cells in lymphocyte cultures stimulated with phytohaemagglutinin or with pokeweed mitogen. MHM6 and AC2 recognized single polypeptides with apparent molecular weights of 45 kd and 80 kd respectively as shown by sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of 125I-labeled cell surface polypeptides immunoprecipitated with these antibodies. These polypeptides were induced on experimentally-infected B cells within 24 h of the expression of the EB virus nuclear antigen, EBNA, at a time known to coincide with the appearance of the lymphocyte-detected membrane antigen, LYDMA. However, saturating concentration of MHM6 and AC2 were unable to protect EB-LCL target cells from lysis by LYDMA-specific cytotoxic T cells in a chromium-release assay.

Animals↗