[CANCER RESEARCH 37, 2291-2296, July 1977]

Cell Surface Markers on Epithelial-Burkitt Hybrid CellsSuperinfected with Epstein-Barr Virus1

Ronald Glaser,2 Gilbert Lenoir, Soldano Ferrane,3 Michele A. Pellegrino, and Guy de-Thé

Department of Microbiology and Specialized Cancer Research Center, The Milton S. Hershey Medical Center, The Pennsylvania State University College ofMedicine, Hershey, Pennsylvania 17033 ¡Ft,G.j; international Agency for Research on Cancer, 150 Cours Albert Thomas, 69008 Lyon, France [G. L., D. G.];and Department of Molecular Immunology, Scripps Clinic and Research Foundation, La Jolla, California 92037 ¡S.F., M. A. P.¡

SUMMARY

Attempts were made to superinfect two epithelial-Burkitthybrid cell lines, designated D98/HR-1 and D98/Raji, withEpstein-Barr virus (EBV) and to investigate the expressionof some cell surface markers, including histocompatibilityantigens, and the presence of B-cell markers, such as receptors for the third complement component and for monkey red blood cells. Successful superinfection of D98/HR-1cells with EBV was made evident by the expression of earlyantigen and, to a lesser extent, virus capsid antigen. Only arare D98/Raji cell was found to be positive for early antigen.The histocompatibility antigens of the parental cell linesD98, HR-1, and Raji were expressed on the surfaces of thehybrid cells. Receptors for third complement components band d were not detected on the hybrid cells or on the D98 orHR-1 cell lines; they were found, however, on the Raji cells,indicating that EBV receptors and complement receptorscan be separated. The significance of the infection of thehybrid cells with EBV and the expression of cell surfacemarkers is described.

INTRODUCTION

We have studied the regulation and expression of theEBV genome in nonlymphoblastoid cell types, i.e., somaticcell hybrids of Burkitt tumor cells (9). With this procedurewe have circumvented the problem of the failure of EBV toinfect various cell types. We have maintained EBV in arepressed state similar to that observed in "nonproducer"

Raji cells for long periods of time and have induced EBV toreplicate and synthesize "complete" virus particles in epi-

1 This study was supported by Contracts N01 CO43296 and N01 CP53516

within the Virus Cancer Program of the National Cancer Institute, NIH,USPHS, and by Grants 1 P30 CA18450, AI13154, CA15038, CA16071, andCA16069. Part of this study was performed at the International Agency forResearch on Cancer, Lyon, France, under the sponsorship of the FogartyInternational Center, the NIH, and the Institut National de la Santéet de laRecherche Médicale, Paris. France. This is Publication 1192 from ScrippsClinic and Research Foundation.

2 Leukemia Society of America Scholar. To whom requests for reprints

should be addressed, at Department of Microbiology, The Milton S. HersheyMedical Center, The Pennsylvania State University, Hershey, Pa. 17033.

3 Recipient of an American Heart Established Investigatorship.4 The abbreviations used are: EBV, Epstein-Barr virus; HLA, histocompati

bility antigens; C3, 3rd complement component; RPMI, Roswell Park Memorial Institute; PCS, fetal calf serum; VCA, virus capsid antigen; PBS, phosphate-buffered saline [0.15 M NaCI. 12 mw Na2HPO, and 1.4 mw KHjPO, (pH7.4)]; ADM, the number of cells (x103//il) required to reduce by 50% thecytotoxic activity of a selected alloantiserum; EAC1-3""', mouse serum defi

cient in the 6th complement component.Received January 26, 1977; accepted April 15. 1977.

thelial hybrid cells after treatment with iododeoxyuridine (6,8, 10).

The detection and characterization of antigenic markersand receptors expressed on the membranes of cells that areinfectable with EBV may enable us to clarify the events inthe infection process and to predict the permissiveness ofother cell types. In this study we investigated cell surfacemarkers expressed on the surfaces of epithelial-Burkitt hybrid cells that are infectable with EBV. Specifically, thesemarkers are HLA and receptors for C3 and monkey RBC,receptors that are specific for B-cells (14).

MATERIALS AND METHODS

Cells. Burkitt's lymphoma cell lines Raji and P3J-HR-1(HR-1) were maintained in RPMI Medium 1640 supplemented with 10% PCS, penicillin (100 units/ml), streptomycin (100 /¿g/ml),and Mycostatin (10 units/ml). Epithelial-Burkitt hybrid cell lines D98/HR-1, clones 1 and 8, and D98/Raji, clones 4 and 16, (7, 9) and the parental D98 cells,which are variants of HeLa cells (5), were maintained inEagle's medium supplemented with 10% PCS, penicillin

(100 units/ml), streptomycin (100 ^g/ml), and Mycostatin(10 units/ml). Cells were grown in plastic tissue cultureflasks (Falcon Plastics, Oxnard, Calif.) at 37°.

Virus. The procedure for preparing EBV concentrates hasbeen described (1). Briefly, HR-1 cells were grown in RPMIMedium 1640 supplemented with 10% PCSat 35°for11 days

without feeding. The cell suspensions were centrifuged at3000 rpm for 20 min, and the supernatants were filteredthrough a 1.2-jum Millipore filter. Virus was precipitatedwith polyethylene glycol and resuspended in 1% of theoriginal volume of RPMI Medium 1640.

Antisera. HLA alloantisera were obtained from the serumbank at the National Institute of Allergy and Infectious Diseases. Antisera to C3 receptors were produced by injecting1 rabbit with 10e Raji cells and reinoculating the animal 14days later. The rabbit was then bled at weekly intervals. Thepresence of antibody to C3 receptors in this serum wasdemonstrated by the ability to inhibit the functional activityof C3 receptors after extensive adsorption with lymphoidcells that do not carry the C3 receptor. Detailed propertiesof the rabbit anti-C3 receptor xenoantisera are describedelsewhere.5

* S. Ferrane and M. A. Pellegrino. Characterization of an Anti-C3 Receptor

Xenoantiserum; submitted for publication.

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ñ.Glaser et al.

Immunofluorescence. Indirect immunofluorescence wasused to detect EBV early antigen or VGA. Glass coverslips ofcell monolayers were washed in PBS, air dried, and fixed inacetone for 10 min at room temperature. Antisera previouslycharacterized for activity against early antigen and VGAwere adsorbed to the cells for 30 min. After being washedwith PBS, the cells were readsorbed with goat anti-human

IgG conjugated to fluorescein isothiocyanate for an additional 30 min. The cells were washed again with PBS,counter-stained with 0.05% Evans blue, mounted in glycerol

on glass slides, and examined with a Leitz microscope withUV.

Superinfection of Burkitt Hybrid Cells with EBV. ClonedD98/HR-1 and D98/Raji cells were grown as monolayers on

glass coverslips. Approximately 24 hr after culturing hadbegun, 0.2 ml of a 10s early antigen-inducing units per ml

suspension EBV (titrated in Raji cells) was added to eachcell culture, and incubation was continued for 1 hr at 37°.After 5 ml of complete Eagle's medium were added to theculture, the cells were incubated for 6 days at 37°and were

then fixed in acetone and examined for EBV early antigenand VGA.

Quantitation of HLA. The amount of HLA on Raji, HR-1,and D98 parent cells and on D98/HR-1 and D98/Raji hybrid

cells was measured by a microquantitative adsorption technique (13). Briefly, HLA alloantisera at a dilution twice theminimal amount required to lyse 95% of the selected targetcells were incubated with cell suspensions at varying concentrations for 60 min at room temperature. The cells werecentrifuged, and the supernatant was examined for residualactivity against selected target cells from 2 unrelated donors of given HLA specificities in a complement-dependent

microcytotoxicity test. The AD5I, parameter was used tocompare results obtained from the different quantitativeadsorptions. Controls of the specificity of the adsorptionassay were performed by adsorptions with sera directedagainst HLA specificities not represented on the cell membrane.

Detection of Receptors for Monkey RBC and C3. Sheeperythrocytes were sensitized with rabbit anti-erythrocyte antibody adjusted to a concentration of 10" cells/ml in Ver-onal-buffered 0.9% NaCI solution and incubated with anequal volume of either rabbit serum deficient in C6 or EAC1-3m" diluted 1:2 in Veronal-buffered 0.9% NaCI solution (4).

Rabbit serum deficient in C6 is immune adherence positive,and EAC1-3"1" is immune adherence negative (4); therefore,

they were used to detect receptors for C3b and C3d, respectively.

The test for rosette formation between lymphoid cells andepithelial cells or epithelial-Burkitt hybrids was performedby incubation of 0.2 ml of cell suspension (106 cells/ml) with0.2 ml of EAC1-3 cells (5 x 107 cells/ml) for 10 min at 37°as

described (4). One drop of the mixture was then put on ahemocytometer and counted for total lymphocytes and ro-setted lymphocytes, i.e., lymphocytes with 3 or more EAC1-

3 attached. For controls, erythrocyte:antibody cells wereused instead of EAC1-3 cells (4). Several double-blind ex

periments were performed to detect C3 receptors. Cellswere coded by Ronald Glaser, shipped to Soldano Ferrone,and tested for C3 receptors; the code was then broken.

We assayed for the presence of receptors for monkey

RBC. Monkey RBC were obtained from a stump-tailed Macaca monkey. The rosette test was performed by centrifuga-

tion of 25 /il of lymphoid cells or epithelial cells, includingD98, D98/HR-1, and D98/Raji (2 x 106 cells/ml), with 25 /xl

of a 1% monkey RBC suspension for 1 min at 400 x g. Thepelleted cells were gently resuspended and then werescored microscopically for rosette formation (14).

RESULTS

Superinfection of Burkitt Hybrid Cells with EBV. D98/HR-1 and D98/Raji cells in culture were infected with EBVprepared from HR-1 cells. Results of these experiments

(Table 1) represent the averages from 6 independent experiments. Some early antigen-positive cells could be detectedby 3 days postinfection; approximately 1% of D98/HR-1

cells were positive for early antigen 6 days after EBV infection (Fig. 1). A very small percentage (0.01%) were VGApositive. Only a rare D98/Raji cell was found to be earlyantigen positive, and no VGA-positive cells were observed.

Under normal conditions the only antigen that can be detected in D98/HR-1 cells is the EBV-associated nuclear anti

gen; early antigen and VGA are repressed. No expression ofEBV-specific antigens was detected in the D98 parent or in

mouse fibroblast (CL1D) cells infected with the same virusstocks. In addition, early antigen was observed in a dividingD98/HR-1 cell in late telophase (Fig. 2), thus confirmingpreviously published data that an early antigen-positive cellcan divide at least 1 time (11).

HR-1 virus-infected cells were fixed and examined by thin-

section electron microscopy. Virus particles could be seenattached to the cell membrane of infected D98/HR-1 cells.No EBV (HR-1 ) virus particles were observed to be adsorbed

to the surface of D98 or D98/Raji cells.Expression of HLA on Burkitt Hybrid Cells. D98/HR-1

clone 1 and 8 and D98/Raji clone 4 and 16 cells expressedHLA from both parental cell lines (Table 2). It could bedetermined that the number of antigenic sites present onthe hybrid cells was similar to that detected on parentallines since similar numbers of both adsorbed equalamounts of HLA alloantisera. D98/HR-1 cells lost the abilityto react with anti-HLA-B12 antiserum, which cross-reactedwith HLA-B5 expressed on the parental line HR-1. In addition, D98/Raji cells acquired the ability to adsorb anti-HLAVictor alloantiserum directed to HLA-B5. Both clones ofD98/Raji cells (including various passage levels) demon-

Table 1Expression of early antigen and VCA in D98 and D98 hybrid cells

after EBV infectionSlides were read by immunofluorescence 6 days postinfection

with early antigen- and VGA-specific antisera. Percentages weredetermined by counting >300 cells in 6 independent experiments.

Presence of EBV-specific markers

CelllineD98CL1D

D98/HR-1D98/RajiEarly

antigen0

01

<0.001VCA000.010

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Cell Surface Markers on EBV-infected Burkitt Hybrid Cells

Table 2Expression of HLA on epithelial-Burkiti hybrids and on their respective parental lines

AD«,

HLA-A HLA-B

line1HR-1D98D98/HR-1Raj

i-D98/Raji21.51.0—1.53

9ND"—

—ND0.72.510

572.0_

__1.5_

__1.5-30.0f8

121250—

--

-——

" AD5I, value greater than 50,000 cells/ml.6 ND, not done.' This range represents the absorbing capacity of 2 clones of D98/Raji at various passage levels

(for explanation see "Results").

strated variability in capacity to adsorb the Victor alloantise-rum, as indicated by the AD.-,0values between 1,500 and30,000 cells//j.l of antiserum. Analysis of the karyotype ofthese hybrids indicated that Chromosome 6, which hasbeen linked to the HLA and C3 receptor phenotype (2, 16),is present in D98/HR-1 (Fig. 3) and D98/Raji (data notshown) cells.

Detection of Receptors for Monkey RBC and C3 on theSurfaces of Burkitt Hybrid Cells. Receptors for C3b andC3d were not detected on D98/HR-1 or D98/Raji cells or onthe respective parental cells, with the exception of Raji cells(Table 3). D98/HR-1 and D98/Raji cells were also unable toadsorb a rabbit xenoantiserum directed to C3 receptors orclosely associated membrane components at the ratio of0.250 x 106 cells//¿Iof antiserum (Chart 1). This antiserumwas adsorbed by Raji cells at the ratio of 30,000 cells//J ofserum and was also adsorbed by other C3 receptor-positivecell lines, e.g., RPMI 6410. Thus, D98/HR-1 cells not expressing detectable levels of C3 receptors were still able tobe superinfected with EBV.

Since the number of early antigen-positive D98/HR-1 cellswas only about 1%, it was possible that the assay used todetect C3 receptors was not sensitive enough to detect thislow level of cells. Therefore, we tested the sensitivity of theassay. Lymphoid cells were mixed with HeLa cells at decreasing concentrations. The percentage of rosette formation was determined (as described in "Materials and Methods") in mixtures in which the lymphoid cells were at 75, 25,

10, 5, 1, and 0% of the cell suspension. Tests were run intriplicate (blind coded) in 2 independent experiments (Chart2). An average of 4.3% rosettes could be detected in whichthe lymphoid cells were 1% of the total cell suspension. Norosettes were observed when HeLa cells were assayedalone, confirming the results obtained with D98 cells.Therefore, the assay used to detect C3 receptors is sensitiveenough to detect 1% early antigen-positive D98/HR-1 cells ifthey also express C3 receptors.

Assays were also performed to determine whether receptors for monkey RBC (associated with B-lymphocytes) wereexpressed on the hybrid cells. It was found that receptorsfor monkey RBC were expressed on the lymphoid cells butnot on the epithelial-Burkitt hybrids or on D98 cells (Table3).

Table 3Expression of C3b, C3d, and monkey RBC receptors on epithelial-

Burkitt hybrids and on their respective parental lines

% rosettes with

CelllineD98

HR-1D98/HR-1

RajiD98/RajiEAC1-30

00

870EAC1-5"0

00

750Monkey

RBC0

910

850

EAC1-5, rabbit serum deficient in C6.

100

80

60

40

20

lililí

À

15 30 60 120

ABSORBING CELLS (103>

240

Chart 1. Adsorbing activity of Raji cells (•)and hybrids D98/Raji (D) andD98/HR-1 (A) for a xenoantiserum to human C3 receptor. Adsorbed serumwas tested for its ability to inhibit the rosette formation between Raji cells andEAC1-31"".A, rosette formation between Raji cells and EAC1-301""in presenceof normal rabbit serum; bars, S.E.

DISCUSSION

We have shown that epithelial-Burkitt hybrid cells (D98/HR-1) can be superinfected with EBV, demonstrated by thesynthesis of EBV early antigen and VGA. The positivity ofthese cells was clear, but the percentages were lower thanthose previously reported when epithelial-Burkitt hybridcells were exposed to iododeoxyuridine and EBV was in-

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R. Glaser et al.

10 20 3O 40 6O

% ROSETTES

80 1OO

Chart 2. Determination of the sensitivity of the rosette formation assayused to detect C3 receptors. Lymphoid cells (C3 receptor positive) wereadded to 2 x 10s HeLa cells (C3 receptor negative) to make final concentrations of 75, 25, 10, 5, and 1% of the cell mixture. The rosette formation assay(described in "Materials and Methods") was used to detect C3 receptors.

duced to replicate (8, 10). In D98/Raji cells, EBV-specificearly antigen could be detected at a very low level. There areseveral EBV genome-positive lymphoblastoid cell lines that,when infected with EBV, demonstrate low levels of earlyantigen that are comparable to those found in D98/HR-1cells (3). The D98 cell line was not superinfectable under thesame conditions and with the same EBV preparation usedfor the susceptible hybrid cells.

Expression of early antigen was recently demonstrated ina Burkitt lymphoblastoid cell in metaphase, suggesting thatan early antigen-positive cell can undergo cell division (11).Four days after EBV infected the culture, we found an earlyantigen-positive D98/HR-1 cell in late telophase. Thus, thesequence of cell division in which early antigen can beexpressed can now be extended to telophase. These dataconfirm the work of Rampar et al. (11). It would seem thenthat a cell that synthesizes at least some component(s) ofearly antigen can divide at least once. What this means interms of the function of early antigen is not clear.

The data obtained in this study suggest that the portion ofthe phenotype of the Burkitt tumor cell that is related to thereceptor for EBV attachment is expressed in the hybridcells. We have also shown that epithelial-Burkitt hybridsexpress HLA present on both parental cell lines. The D98/HR-1 hybrids lose the ability to adsorb cross-reacting antibodies to HLA-B12, which is expressed on both parental celllines. In contrast, D98/Raji cells can adsorb an anti-HLA-B5alloantiserum although neither of the parental cells canreact with it. Adsorption of Victor serum is likely to reflect across-reaction between HLA-B5 and HLA-W35; the latterhas reportedly been expressed on D98 cells (12). Changesof reactivity of hybrid cells with antisera to cross-reactingHLA specificities may reflect conformational changes ofantigenic moieties inserted in the cell membrane and/orexposure of concealed membrane determinants as a resultof alterations in membrane biosynthesis.

Two points regarding the expression of C3 receptors onhybrid cells deserve comment. Firstly, D98/HR-1 hybridcells could be infected with EBV although C3 receptorswere not detected. It has recently been suggested that C3receptors may act as or be associated with EBV receptors(17). Our findings indicate that infection with EBV can occurwithout detectable C3 receptors. We have also shown thatthe procedure used to measure C3 receptors is sensitiveenough to detect 1% of cells with C3 receptors, discountingthe possibility that we would not detect the 1% early antigen-positive cells if they possessed the C3 receptor.

Secondly, the D98/Raji hybrids do not express C3 receptors although the Raji parental line carries C3 receptors.The lack of expression of C3 receptors on D98/Raji cells isreminiscent of the lack of secretion of a-fetoprotein andalbumin from hybrids of mouse hepatoma cells (which secrete these proteins) and mouse or rat fibroblasts (which donot secrete them) (15). Recently, a synteny between HLA,C3 receptors, and Chromosome 6 has been shown (2, 16).Since the hybrid cells carry HLA antigens and contain Chromosome 6, their lack of C3 receptors cannot result from aloss of the chromosome carrying the genetic informationfor C3. The following possibilities can be evisioned for ourexperimental finding: (a) the C3 receptor is present on thecell membrane but cannot be detected because it is coatedby a "masking" substance; or (b) the expression of C3

receptors is blocked at the level of synthesis, reflectingsome interaction between the 2 genomes present in thehybrid nucleus, and/or at insertion in cell membranes. Thelatter possibility seems most probable since cells such asD98 may carry HLA although not expressing C3 receptoractivity. This explanation would also explain the resultsobtained with the receptors for monkey RBC. These receptors were expressed on the surface of both Burkitt lymphoblastoid parental cell lines but not on either D98/HR-1 orD98/Raji cell lines.

We have shown that certain epithelial-Burkitt hybrid cellscan be infected with EBV and that these cells express theHLA of both parental cell lines but not of C3 receptors. Theavailability of hybrid cells that lack cell surface markersexpressed on 1 parental line is useful to define the geneticcontrol and the regulatory mechanisms involved in theirexpression and to investigate their possible function in virusinfection.

ACKNOWLEDGMENTS

The authors acknowledge the excellent technical assistance of M. F.Lavoue, M. C. Favre, C. Piccoli, and G. Baughman. We also thank M.Vuilloume for electron microscopy data, R. Ladda and Y. Dobelle for helpfuldiscussion and chromosome analysis, and F. Rapp for constructive criticismof the manuscript.

REFERENCES

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2. Curry, R. A., Dierich, M. P., Pellegrino, M. A., and Hoch, J. A. Evidencefor Linkage between HLA Antigens and Receptors for Complement Components C3b and C3d in Man-Mouse Hybrids. Immunogenetics, 3. 465-472, 1976.

3. Diehl, V., Wolf, H., Schulte-Holthausen, H., and zur Hausen, H. Re-exposure of Human Lymphoblastoid Cell Lines to Epstein-Barr Virus.

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Intern. J. Cancer, 70: 641-651, 1972. 124. Dierich, M. P., Pellegrino, M. A., Ferrane, S., and Reisfeld, R. A. Evalua

tion of C3 Receptors on Lymphoid Cells with Different ComplementSources. J. Immunol., 112: 1766-1773, 1974. 13.

5. Gartler, S. M. Apparent HeLa Cell Contamination of Human HeteroploidCell Lines. Nature, 277: 750-751, 1968.

6. Glaser, R., Farrugia, R., and Brown, N. Effect of the Host Cell on the 14.Maintenance and Replication of Epstein-Barr Virus. Virology, 69: 132-142, 1976.

7. Glaser, R., and Nonoyama, M. Host Cell Regulation of Induction of 15Epstein-Barr Virus. J. Virol., 14: 174-176, 1974.

8. Glaser, R., Nonoyama, M., Decker, B., and Rapp, F. Synthesis of Ep- 16.stein-Barr Virus Antigens and DMA in Activated Burkitt Somatic CellHybrids. Virology, 55:62-69, 1973.

9. Glaser, R., and O'Neill, F. J. Hybridization of Burkitt LymphoblastoidCells. Science, 176: 1245-1247, 1972.

10. Glaser, R., and Rapp, F. Rescue of Epstein-Barr Virus from Somatic Cell 17Hybrids of Burkitt Lymphoblastoid Cells. J. Virol., 70: 288-296, 1972.

11. Hampar, B., Derge, J.G., Nonoyama, M., Chang, S-Y., Tagamets, M. A.,and Showalter, S. D. Programming of Events in Epstein-Barr Virus-Activated Cells by 5-iododeoxyuridine. Virology, 62: 71-89, 1974.

Kennett, R. H., Hampshire, B., Bengtsson, B., and Bodmer, W. F.Expression and Segregation of HL-A Antigens in D98/AH-2 by Lymphocyte and Fibroblast Hybrids. Tissue Antigens, 6: 80-92, 1975.Pellegrino, M. A., Ferrane, S., and Pellegrino, A. A Simple Microabsorption Technique for HL-A Typing. Proc. Soc. Exptl. Biol. Med., 739:484-488,1972.Pellegrino, M. A., Ferrane, S., and Theofilopoulos, A. N. Rosette Formation of Human Lymphoid Cells with Monkey Red Blood Cells. J. Immunol., 775: 1065-1071, 1975.Szpirer, J., and Szpirer, C. The Control of Serum Protein Synthesis inHepatoma-Fibroblast Hybrids. Cell, 6: 53-60, 1975.Van Someren, H., Westerveld, A., Hagemeijer, A., Mees, J. R., Khan, P.M., and Zaalberg, 0. B. Human Antigen and Enzyme Markers in Man-Chinese Hamster Somatic Cell Hybrids: Evidence for Synteny betweenthe HL-A, PGM:,, ME, and IPO-B Loci. Proc. Nati. Acad. Sei. U. S., 77:962-965, 1974.Yefenof, E., Klein, G., Jondal, M., and Oldstone, M. B. A. SurfaceMarkers on Human B and T Lymphocytes. IX. Two-Color Immunofluores-cence Studies on the Association between EBV Receptors and Complement Receptors on the Surface of Lymphoid Cell Lines. Intern. J. Cancer, 77: 693-700, 1976.

Fig. 1. Immunofluorescence photomicrograph of D98/HR-1 clone 1 cells infected with EBV derived from HR-1 cells 6 days postinfection, showing earlyantigen. Note cytoplasmic fluorescence. Approximately x 1125.

Fig. 2. Immunofluorescence photomicrograph of a D98/HR-1 clone 1 cell infected with EBV derived from HR-1 cells 4 days postinfection. Note that theearly antigen-positive cell is in late telophase. Approximately x 1700.

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Fig. 3. Photograph of trypsin-banded metaphase chromosomes of D98/HR-1 cells. Note 2 No. 6 chromosomes (arrows). D98/Raji cells also contain No. 6chromosomes (not shown). Approximately * 6000.

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1977;37:2291-2296. Cancer Res   Ronald Glaser, Gilbert Lenoir, Soldano Ferrone, et al.   Superinfected with Epstein-Barr VirusCell Surface Markers on Epithelial-Burkitt Hybrid Cells

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