Detection and Localization of Epstein-Barr Virus-Associated Early Antigens in Single Cells by Autoradiography Using 125I-Labeled Antibodies

ABSTRACT A fundamental problem in studying the latent-to-lytic switch of Epstein-Barr virus (EBV) and the viral lytic cycle itself is the lack of a culture system fully permissive to lytic cycle induction. Strategies to target EBV-positive tumors by inducing the viral lytic cycle with chemical agents are hindered by inefficient responses to stimuli. In vitro, even in the most susceptible cell lines, more than 50% of cells latently infected with EBV are refractory to induction of the lytic cycle. The mechanisms underlying the refractory state are not understood. We separated lytic from refractory Burkitt lymphoma-derived HH514-16 cells after treatment with an HDAC inhibitor, sodium butyrate. Both refractory- and lytic-cell populations responded to the inducing stimulus by hyperacetylation of histone H3. However, analysis of host cell gene expression showed that specific cellular transcripts Stat3, Fos, and interleukin-8 (IL-8) were preferentially upregulated in the refractory-cell population, while IL-6 was upregulated in the lytic population. STAT3 protein levels were also substantially increased in refractory cells relative to untreated or lytic cells. This increase in de novo expression resulted primarily in unphosphorylated STAT3. Examination of single cells revealed that high levels of STAT3 were strongly associated with the refractory state. The refractory state is manifest in a unique subpopulation of cells that exhibits different cellular responses than do lytic cells exposed to the same stimulus. Identifying characteristics of cells refractory to lytic induction relative to cells that undergo lytic activation will be an important step in developing a better understanding of the regulation of the EBV latent to lytic switch.

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A microassay for quantitatively detecting the epstein-barr virus receptor on single cells utilizing flow cytometry

Journal of Virological Methods ◽

10.1016/0166-0934(81)90047-1 ◽

1981 ◽

Vol 3 (3) ◽

pp. 127-136 ◽

Cited By ~ 3

Author(s):

Alan Wells ◽

Harald Steen ◽

Sem Saeland ◽

Tore Godal ◽

George Klein

Keyword(s):

Flow Cytometry ◽

Epstein Barr Virus ◽

Single Cells ◽

Virus Receptor ◽

Barr Virus ◽

Epstein Barr

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A Quantitative Ultrastructural Study of Peripheral Blood Lymphocytes Containing Parallel Tubular Arrays in Epstein-Barr Virus and Cytomegalo-Virus Mononucleosis

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100098769 ◽

1981 ◽

Vol 39 ◽

pp. 454-455

Author(s):

C. M. Payne ◽

P. M. Tennican

Keyword(s):

Peripheral Blood ◽

Lymphocyte Count ◽

Epstein Barr Virus ◽

Absolute Lymphocyte Count ◽

Peripheral Blood Lymphocytes ◽

Clinical State ◽

Barr Virus ◽

The Absolute ◽

Epstein Barr ◽

Tubular Arrays

In the normal peripheral circulation there exists a sub-population of lymphocytes which is ultrastructurally distinct. This lymphocyte is identified under the electron microscope by the presence of cytoplasmic microtubular-like inclusions called parallel tubular arrays (PTA) (Figure 1), and contains Fc-receptors for cytophilic antibody. In this study, lymphocytes containing PTA (PTA-lymphocytes) were quantitated from serial peripheral blood specimens obtained from two patients with Epstein -Barr Virus mononucleosis and two patients with cytomegalovirus mononucleosis. This data was then correlated with the clinical state of the patient.It was determined that both the percentage and absolute number of PTA- lymphocytes was highest during the acute phase of the illness. In follow-up specimens, three of the four patients' absolute lymphocyte count fell to within normal limits before the absolute PTA-lymphocyte count.In one patient who was followed for almost a year, the absolute PTA- lymphocyte count was consistently elevated (Figure 2). The estimation of absolute PTA-lymphocyte counts was determined to be valid after a morphometric analysis of the cellular areas occupied by PTA during the acute and convalescent phases of the disease revealed no statistical differences.

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Studies of Epstein-Barr Virus Antigens Using Immunoperoxidase and Immunofluorescence Techniques

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010011533x ◽

1975 ◽

Vol 33 ◽

pp. 342-343

Author(s):

R. Stephens ◽

K. Traul ◽

D. Woolf ◽

P. Gaudreau

Keyword(s):

Electron Microscopy ◽

Nasopharyngeal Carcinoma ◽

Epstein Barr Virus ◽

Barr Virus ◽

If Technique ◽

Infected Cells ◽

Virus Antigens ◽

Epstein Barr ◽

Virus Capsid Antigen ◽

Antigen Reactivity

A number of antigens have been found associated with persistent EBV infections of lymphoblastoid cells. Identification and localization of these antigens were principally by immunofluorescence (IF) techniques using sera from patients with nasopharyngeal carcinoma (NPC), Burkitt lymphoma (BL), and infectious mononucleosis (IM). Our study was mainly with three of the EBV related antigens, a) virus capsid antigen (VCA), b) membrane antigen (MA), and c) early antigens (EA) using immunoperoxidase (IP) techniques with electron microscopy (EM) to elucidate the sites of reactivity with EBV and EBV infected cells.Prior to labeling with horseradish peroxidase (HRP), sera from NPC, IM, and BL cases were characterized for various reactivities by the indirect IF technique. Modifications of the direct IP procedure described by Shabo and the indirect IP procedure of Leduc were made to enhance penetration of the cells and preservation of antigen reactivity.

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