EB病毒相关胃癌组织中病毒基因的表达

背景与目的：EB病毒(Epstein—Barr virus,EBV)与多种恶性肿瘤的发生有关,在鼻咽癌及淋巴瘤等组织中EBV的存在形式和表达已有报道,研究表明不同肿瘤组织中EBV的存在形式和表达不同,在胃癌组织中EBV某些基因尤其是裂解性基因的表达情况报道较少。为明确胃癌组织中EBV潜伏性基因和裂解性基因的表达情况,本研究从mRNA水平检测胃癌组织中EBV潜伏感染和裂解感染相关基因的表达,从分子水平探讨EBV编码基因与胃癌发生发展的关系。方法：应用PCR—Southern杂交检测185例胃癌及其相应癌旁组织中特异性EBVDNA片段,进一步用原位杂交(ISH)技术检测PCR阳性标本EBV编码小RNA1(EBERl)的表达,癌细胞EBERl阳性者确定为EBV相关胃癌(EBV—associated gastric carcinoma,EBVaGC)。采用RT-PCR和Southern杂交技术检测EBVaGCs组织中EBV核抗原(EBNAs)基因转录启动子(Qp、Cp和Wp)、潜伏性基因(EBNA1、EBNA2基因,潜伏膜蛋白LMP1、LMP2A和LMP2B基因)和裂解性基因(即刻早期基因BZLF1和BRLF1,早期基因BARF1和BHRF1,晚期基因BcLF1和BLLF1)的表达。结果：185例胃癌标本EBV阳性率为7．03％(13／185),癌旁组织均为阴性。13例EBVaGCs组织中均检测到启动子Qp mRNA,而Wp和Cp mRNA则为阴性。潜伏性基因中EBNA1 mRNA均阳性(13／13),而EBNA2、LMP1和LMP2B均未见表达,5例标本(5／13)检测到LMP2A mRNA。裂解性基因中BcLF1有7例(7／13)表达阳性,2例(2／13)BHRF1表达阳性,6例标本(6／13)检测到BZLF1 mRNA,BARF1亦有6例(6／13)表达阳性,而BRLF1和BLLF1mRNA均为阴性。结论：EBVaGC中EBV潜伏类型为Ⅰ型或介于Ⅰ和Ⅱ型之间的独特类型;EBVaGC组织中部分裂解性基因表达阳性,部分胃癌存在BARF1和BHRF1表达,其在胃癌中所起的作用有待进一步研究。     

EB病毒相关胃癌与mdm2，p53及p21基因异常的研究

 目的 探讨p53基因突变以及mdm2，p53和p21蛋白表达异常在EBV相关胃癌（EBVaGC）发生发展中的作用。方法 应用免疫组化技术检测13例EBVaGC、45例临床病理资料与之匹配的EBV阴性胃癌（EBVnGC）组织中 mdm2，p53和p21蛋白的表达；PCR-SSCP银染技术结合DNA序列分析检测p53基因exon 5~8突变；RT-PCR检测EBV相关基因的表达。结果 E-BVaGC组与EBVnGC组相比，两组间mdm2，p53和p21蛋白的阳性检出率差异无统计学意义（P = 0.830 0；P = 0.791 2；P = 0.353 1），但EBVaGC组p53蛋白过表达率（15.38 %）明显低于EBVnGC组（57.78 %），两组间差异有统计学意义（P = 0.008 5）。mdm2蛋白阳性表达与p53蛋白过表达呈显著正相关（P = 0.000 8，r = 0.439 1）；p21与p53蛋白共同表达率较高，但经计数资料相关性统计学分析表明两者无显著相关性（P = 0.2501，r = 0.202 5）。2例EBVnGC检测到p53基因突变，突变均位于exon 5，13例E-BVaGC和58例相应癌旁组织均未检测到p53基因突变。13例EBVaGC核抗原基因EBNA1均为阳性，潜伏膜蛋白基因LMP1均为阴性，即刻早期基因BZLF1，早期基因BARF1和BHRF1阳性率分别为46.15 %（6／13），46.15 %（6／13）和15.38 %（2／13），三者与EBVaGC组织中mdm2，p21和p53蛋白的表达均无显著相关性（P＞0.05）。 结论 EBV感染以及mdm2，p53和p21蛋白表达异常与胃癌发生有关； p53基因突变可能并非胃癌组织中p53蛋白异常累积的主要原因；胃癌组织中EBV感染与p53蛋白的异常表达有关，而与mdm2和p21蛋白的异常表达以及p53基因突变无显著相关性。     

EBV相关胃癌的临床病理学分析

目的：探讨Epstein-Barr病毒（EBV）感染与胃癌发生的关系，以及EBV相关胃癌和EBV阴性胃癌在临床病理资料方面的异同。方法：应用PCR-Southern技术检测185例胃癌及相应癌旁组织EBVDNA，并用原位杂交技术检测PCR阳性标本EBV编码的小RNA（EBERs），证实EBV感染的存在。结果：185例胃癌组织标本EBV阳性率为7．03％（13／185），而相应癌旁组织均为阴性，两种组织中EBV阳性率差异有统计学意义，X^2=11．0709，P=0．0009。结论：EBV相关胃癌和EBV阴性胃癌在性别、年龄、肿瘤大小、组织学类型、分化程度、淋巴结转移以及临床病理分期等方面差异均无统计学意义，但两组间肿瘤发生的部位差异有统计学意义。     

EBV相关胃癌的临床病理学分析

目的探讨Epstein-Barr病毒(EBV)感染与胃癌发生的关系,以及EBV相关胃癌和EBV阴性胃癌在临床病理资料方面的异同。方法应用PCR-Southern技术检测185例胃癌及相应癌旁组织EBV DNA,并用原位杂交技术检测PCR阳性标本EBV编码的小RNA(EBERs),证实EBV感染的存在。结果185例胃癌组织标本EBV阳性率为7·03%(13/185),而相应癌旁组织均为阴性,两种组织中EBV阳性率差异有统计学意义,χ2=11·0709,P=0·0009。结论EBV相关胃癌和EBV阴性胃癌在性别、年龄、肿瘤大小、组织学类型、分化程度、淋巴结转移以及临床病理分期等方面差异均无统计学意义,但两组间肿瘤发生的部位差异有统计学意义。     

缺氧诱导因子-1α在胃癌中的表达及其与肿瘤血管生成及细胞增殖的关系

目的检测胃癌组织中缺氧诱导因子-1α（HIF-1α）、血管内皮生长因子（VEGF）及增殖细胞核抗原（PCNA）,并探讨其临床意义。方法采用免疫组织化学sP法检测正常胃黏膜及胃癌中的HIF-1α、VEGF、及PCNA。结果腺癌中HIF-1α阳性表达率显著高于正常组织（X^2=9．35,P〈0．05）,腺癌中VEGF阳性表达率显著高于正常组织（X^2=11．94,P〈0．05）,腺癌中PCNA高增殖活性表达率显著高于正常组织（X^2=8.31,P〈0．05）;有淋巴结转移者HIF-1α阳性表达率显著高于无淋巴结转移者（X^2=6．21,P〈0．05）,随着浸润深度的增加,HIF-1α阳性表达率逐渐增加。HIF-1α阳性表达率与胃癌肿瘤大小、分化程度无关。有淋巴结转移者VEGF阳性表达率显著高于无淋巴结转移者（X^2=5．99,P〈0．05）,随着浸润深度的增加,VEGF阳性表达率逐渐增加。VEGF阳性表达率与胃癌肿瘤大小、分化程度无关。有淋巴结转移者PCNA阳性表达率显著高于无淋巴结转移者（X^2=5．19,P〈0．05）。随着浸润深度的增加,PCNA阳性表达率逐渐增加。PCNA阳性表达率与胃癌肿瘤大小、分化程度无关。在胃癌中HIF表达与VEGF呈正相关（X^2=8．42,P〈0．05）,HIF表达与PCNA呈正相关（X^2=7．35,P〈0．05）。结论HIF-1α通过促进肿瘤血管形成及细胞增殖,在胃癌的发生、发展中发挥重要作用：     

EBV相关胃癌与P53基因异常的研究

[目的]探讨EBV感染和p53基因异常在胃癌发生发展中的病因学作用.[方法]应用免疫组化技术检测13例EBV相关胃癌(EBV associated gastric carcinoma,EBVaGC),45例临床指标与之匹配的EBV阴性胃癌(EBV negative gastric carcinoma,EBVnGC)以及58例相应癌旁组织中p53蛋白的表达;PCR-SSCP银染技术检测p53基因exon 5～8突变情况.[结果]①胃癌组p53蛋白阳性率为86.2%(50/58),而相应癌旁组织均为阴性,胃癌组p53蛋白的阳性率明显高于癌旁组织组,两组间有极显著性差异(P=0.0000).②EBVnGC p53蛋白阳性率为86.7%(39/45),过表达率为57.8%(26/45);EBVaGC p53蛋白阳性率为84.6%(11/13),过表达率仅为15.4%(2/13).两组间p53蛋白的阳性检出率无明显差异(P=0.7912),但EBVaGC组p53蛋白过表达率明显低于EBVnGC组,两组间有显著性差异(P=0.0085).③2例EBVnGC检测到p53基因突变,突变均位于exon 5,13例EBVaGC和58例相应癌旁组织均未检测到p53基因突变.[结论]p53基因异常与胃癌的发生密切相关,EBVaGC组织中存在p53蛋白的表达和过表达,但p53蛋白的异常累积可能并非p53基因突变所致.     

Epstein—Barr病毒在肝细胞癌发生中的作用探讨

目的：了解肝细胞癌(hepatocellular carcinoma，HCC)患者的EpsteinBarr病毒(EBV)感染情况，探讨EBV与肝炎病毒有无协同致癌作用。方法：研究组为78例HCC石蜡标本，对照组为26例非癌症肝组织标本。用PCR检测EBV，DNA(BarnHl W，LMP1)、HBV DNA(S基因、X基因)，用RT—PCR检测HCV RNA和HDV RNA，用免疫组化检测EBV(LMP1)、HBV(HBsAg、HbcAg)和HCV。结果：EBV DNA在HCC组阳性率高于对照组(28．2％vs8．0％)，X^2=4．622，P=0．032；HBV DNA在HCC组阳性率高于对照组(56．4％vs23．1％)，X^2=8．681，P=0．008；EBV与HBV在HCC组无相关关系，X^2=0．835，P=0．375。HCV RNA、HDV RNA在18例HCC中阳性分别为1和0例。免疫细化测EBV在HCC组阳性率高于对照细(32．1％vs2．5％)，X^2=6．02，P=0．012；HBV在HCC组阳性率高于对照细(57．7％vs5．1％)，X^2=10．03。P=0．001。结论：EBV在HCC发生中可能起作用，与HBV无明显协同致癌作用;HCV、HDV检出率不高，与EBV关系未能确定。     

乳腺肿瘤组织中EGFR与EGFRvⅢ的表达及临床意义

目的：探讨EGFR和EGFRvⅢ在人乳腺癌及乳腺良性增生性疾病中的表达及临床意义。方法：应用Supervision^TM免疫组化方法检测30例乳腺良性增生性病变、病变旁及55例乳腺癌、癌旁和转移淋巴结组织的EGFR、EGFRvⅢ的表达。结果：EGFR在乳腺癌表达72.7％，癌旁29.1％，转移淋巴结58．3％（X^2=5.326，P=0.021）；EGFRvⅢ在乳腺癌表达52.7％，癌旁1．8％，转移淋巴结45．8％（X^2=4．718，P=0．030）。EGFR表达与病理分级（X^2=11.840，P=0．002）、转移淋巴结数目（X^2=13．715，P=0．000）、ER阴性（X^2=10．785，P=0．007）、PR阴性（X^2=4.727，P=0．031）相关；EGFRvⅢ与病理分级（X^2=5．280，P=0．024）、有无远处转移（X^2=5．612，P=0.019）、淋巴结转移数目（X^2=4．464，P=0.038）、ER阴性（X^2=3．960，P=0．049）、PR阴性（X^2=17．517，P=0．000）和c-erbB-2阳性（X^2=7．424，P=0．005）相关。结论：EGFR、EGFRvⅢ过度表达可能与乳腺癌发生发展密切相关，EGFRvⅢ有望作为预测乳腺癌临床预后候选基因之一。     

胃癌组织中p53基因突变及p53和mdm2蛋白表达的研究

目的：探讨mdm2和p53基因异常在胃癌发生发展中的作用以及两者相关性。方法：应用免疫组化技术检测58例胃癌组织以及相应癌旁组织中mdm2和p53蛋白的表达；PCR-SSCP银染技术检测p53基因exon5～8突变情况。结果：胃癌组p53和mdm2蛋白阳性率分别为86．21％（50／58）和29．31％（17／58），癌旁组织组p53和mdm2蛋白均为阴性，胃癌组p53和mdm2蛋白阳性率明显高于癌旁组织组，两两间差异有统计学意义，P=0．0000、P=0．0001。胃癌组织中p53和mdm2蛋白表达率与肿瘤大小、组织学类型、分化程度、淋巴结转移以及患者年龄等无显著相关性，P〉0．05。mdm2蛋白阳性表达与p53蛋白过表达呈显著正相关,X^2=11．1839，P=0．0008，r=0．4391。2例胃癌组织检测到p53基因突变，突变均位于exon5，58例相应癌旁组织均禾检测到p53基因突变。结论：mdm2和p53蛋白异常表达与胃癌发生有关，p53基因突变可能并非胃癌组织中p53蛋白异常累积的主要原因，mdm2蛋白在胃癌发生发展中的作用可能与p53蛋白密切相关。     

胃癌组织幽门螺杆菌感染与EB病毒潜伏膜蛋白表达的相关性

目的：观察胃癌组织中幽门螺杆菌（H．pylori）感染和EB病毒潜伏膜蛋白（EBV—LMP）的同步表达情况，探讨两者在胃癌中的相互关系。方法：80例正常胃黏膜组织和97例胃癌组织，利用HE染色对所取标本进行组织病理学诊断，EBV—LMP测定采用免疫组化sP法，H．pylori感染判定采用HE，H．pylofi—DNA PCR和血清ELISA法测定IgG抗体三种方法。结果：正常胃黏膜组织未见EBV—IMP的表达，胃癌黏膜组织EBV—LMP表达阳性率为7．2％（7／97），胃癌组织EBV—LMP表达明显高于正常胃黏膜组织，差异有统计学意义（P〈0．05）。本组病例中H．pylofi感染的胃癌黏膜组织中EBV—LMP的阳性表达率为13．5％（7／52），无H．pylori感染的胃癌组织EBV—LMP的阳性表达率为0，H．pylori感染的胃癌组织EBV—LMP表达明显高于无H．pylofi感染的胃癌组织，差异有统计学意义（P〈0．05）。结论：正常胃黏膜组织无EBV—LMP的表达，胃癌组织中EBV—LMP表达明显高于正常胃黏膜组织，H．pylofi感染胃癌组织比无H．pylori感染胃癌组织EBV—LMP呈现高表达，H．pylofi感染和EB病毒感染在胃癌的发生和发展过程中可能具有更加深在的相互关系。     

Identification of EBV transforming genes by recombinant EBV technology

Epstein-Barr virus (EBV) is able to infect primary B-lymphocytes but usually does not proceed to replicate more virions. Instead, EBV persists as an incomplete virus and expresses 12 gene products that transform the growth of these cells into continuously proliferating lymphoblastoid cell lines. Because EBV is associated with several human malignancies, there is intense interest in delineating the molecular functions of these EBV gene products in transformation. This review focuses on the recombinant EBV technologies that have been developed to introduce specific mutations into EBV and test the functions of these EBV genes in primary B-lymphocyte growth transformation.     

EBV conference establishes goals for defining disease-related EBV subtypes for vaccine development

Epstein-Barr virus （EBV）, a γ-herpesvirus, was identified more than four decades ago. It is a ubiquitous infectious agent that infects more than 90% of the world＇s adult population. Primary EBV infection in most individuals is asymptomatic or shows symptoms similar to other childhood illnesses. However, primary EBV infection in adolescents can cause infectious mononucleosis with symptoms that include sore throat, fever, and swollen glands. EBV infection can also affect the liver, heart, and spleen, but even in severe eases, it is rarely fatal.     

EBVDNA和EBV抗体指标早期诊断鼻咽癌方案的筛选

目的:探讨EB病毒DNA及EB病毒抗体在鼻咽癌联合诊断中的应用价值.方法:对81例未经治疗的病理确诊鼻咽癌怠者和89例健康体检者分别采用实时荧光定量PCR测定EB病毒DNA,计算病毒拷贝数,采用间接免疫酶法测定EBV VCA/IgA和EMIgA,采用酶联吸附试验(ELISA法)检测EBNA1/IgG、EBNA1/IgA以及ZTA/IgG.通过ROC曲线评价各指标单独和联合诊断鼻咽癌的效果.结果:分析发现VCA/IgA、EBV DNA和EBNA1/IgA 3个指标具有较高的灵敏度,分别为95.06%、83.95%和81.63%.而EA/IgA、EBNA1/IgG和ZTA/IgG则具有较高的特异度,分别为94.38%、95.51%和89.89%.EBNA1/IgA与VCM/IgA并联时灵敏度为100%,特异度为84.27%.结论:EB-NA1/IgA与VCA/IgA并联时灵敏度和特异度都在比较高的水平,可作为鼻咽癌早期筛查的选择方案.     

Integrated Epstein-Barr virus (EBV) and chromosomal abnormality in chronic active EBV infection

In order to examine the role of Epstein-Barr virus (EBV) in the pathogenesis of chronic active EBV infection (CAEBV), we investigated whether or not EBV integration into the human genome is associated with any chromosonal abnormality. We therefore analyzed 4 cases of CAEBV: 2 cases showed a normal karyotype, while one had an oligo-clonal 6th chromosomal abnormality and the fourth had a clonal 6th deletion (q15q23). In addition, the case with an oligo-clonal abnormality also had oligo-clonal EBV terminal repeat (TR) bands, while the case with a clonal abnormality showed a clonal TR band. In contrast, the 2 cases with a normal karyotype showed no clonal band. Two-color fluorescence in situ hybridization (FISH) was used to detect the integrated EBV and the 6th chromosomal site. The presence of integrated EBV into the 6th chromosome was not frequent in the 2 cases with a normal karyotype, but it was statistically frequent in the case with an oligo-clonal 6th abnormality. In the case with a clonal 6th deletion, integration in the 6th chromosome was also slightly higher than that in the other chromosomes. In CAEBV, integrated EBV might thus be associated both with chromosomal abnormality and with pathogenesis. Int. J. Cancer 71: 943-947, 1997. © 1997 Wiley-Liss Inc.     

Recovery of transforming EBV from non-producer cells after superinfection with non-transforming P3HR-1 EBV.

Cells of the Raji and NC37 lines can be induced by chemical inducers, such as BrdUrd and IdUrd, or the tumor-promoter TPA to EA-expression only, but do not reveal any VCA synthesis. After superinfection by nontransforming P3HR-1 EBV, however, a varying percentage of the cell population shows VCA synthesis and releases infectious viral particles. The recovered virus differs biologically from P3HR-1 EBV since it transforms human umbilical cord blood lymphocytes into EBNA-positive lymphoblastoid cell lines. Cells of these established lines are susceptible to renewed infection by P3HR-1 EBV which results in EA induction and VCA synthesis. Only cells of one line, NC37-R1, spontaneously produce VCA and EBV particles, which reveal transforming properties and do not induce EA upon superinfection of Raji cells. Infection of P3HR-1 EBV-converted BJA-B cells also leads to EA and VCA induction and the release of viral particles. In contrast to particles recovered from Raji and NC37 cells, no transforming activity was detectable in these virus preparations. According to these data, we propose that viral genomes persisting within Raji and NC37 cells are defective and become complemented by the superinfecting P3HR-1 virus.     

EBV infection of mitogen-stimulated human B lymphocytes

B lymphocytes from human blood were treated with the mitogen Protein A (Staph, aureus), which induces DNA synthesis in, as well as differeniation of, the B lymphocytes. The cells were subsequently exposed to Epstein-Barr virus (EBV). EBV-infected and mitogen-stimulated cells were detected simultaneously, by using a combination of immunofluorescence (for EBNA) and autoradiography (for DNA-synthesis). In the initial phase (1-2 days) after addition of the mitogen, stimulated cells were as susceptible to infection as resting cells. Thereafter, their susceptibility decreased. This suggests that, although initiation of DNA-synthesis does not seem to limit sensitivity to the viral infection, differentiation of the B cells might do so. The intensity and pattern of EBNA-staining in prestimulated cells different from that of the controls.     

The characteristics of Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma: comparison between EBV(+) and EBV(-) cases in Japanese population.

We have investigated 114 cases with diffuse large B-cell lymphoma (DLBCL) to clarify the characteristics of DLBCL with Epstein-Barr virus (EBV) infection. Thirteen cases (11.4%) showed EBV-encoded RNA 1 (EBER1) signals by RNA in situ hybridization. EBV-encoded latent membrane protein 1 (LMP1) and EBV-encoded nuclear antigen 2 (EBNA2) were expressed in 11 and 4 cases, respectively. Expression of CD30, Bcl-6 and immunoglobulin (Ig) was found in 92%, 31% and 23% with EBV(+) DLBCL, and in 15%, 79% and 82% with EBV(-) DLBCL, respectively. The sequence of rearranged Ig heavy chain (IgH) variable (V) region gene was analyzed in 5 cases with EBV(+) DLBCL and 61 cases with EBV(-) DLBCL. Somatic mutation was found in all cases except one with EBV(-) DLBCL. Average mutation frequency was 9.6% in EBV(+) DLBCL vs. 11.5% in EBV(-) DLBCL. The rates of replacement mutation vs. silent mutation (R / S values) in complementarity determining region II and framework region III were 2.7 and 1.5 in EBV(+) DLBCL, 2.6 and 1.4 in EBV(-) DLBCL. Crippling mutation generating a stop codon was found in 2 of 5 cases (40%) with EBV(+) DLBCL, but none of 61 cases (0%) with EBV(-) DLBCL. These findings suggest that EBV(+) DLBCL and EBV(-) DLBCL were both derived from germinal center (GC) or post-GC B cells, and EBV(+) DLBCL frequently have a non-functional IgH gene owing to crippling mutation.     

The interaction of non-transforming Epstein-Barr virus (EBV) with cell-associated EBV DNA in superinfected lymphoblastoid cell lines

Two human lymphoblastoid cell lines were established by the transformation of human cord-blood lymphocytes with transforming Epstein-Barr virus (EBV). One cell line (HLB-R1) was established with EBV obtained after the superinfection of Raji cells with HR-1 EBV and the other (HLB-Bl) was established from B95-8 EBV-infected human cord-blood lymphocytes. Both the HLB-R1 and HLB-B1 lines were susceptible to superinfection with HR-1 EBV. We found that EBV DNA was replicated in the superinfected cell lines and that transforming EBV was produced in both the HLB-B1 and HLB-R1 cells. The average titer of transforming EBV obtained in the HR-1 EBV superinfected HLB-B1 and HLB-R1 cell lines was 10(4) transforming units (TU)/ml, whereas the average titers of transforming EBV obtained by the superinfection of Raji cells was 10(1) TU/ml. Epstein-Barr virus capable of inducing early antigen (EA) in superinfected Raji cells (lytic virus) was not detected in any transforming virus preparation. Restriction enzyme digestion patterns of virus DNA isolated from HR-1 and B95-8 cells, as well as from superinfected cells, were compared. The EBV DNA that was replicated in the superinfected HLB-R1 and HLB-B1 cell lines showed a more complex pattern. Our data suggest that recombination between input HR-1 EBV DNA and latent cell-associated EBV DNA occurs. Presumably this recombination results in a change in the biological properties of the newly synthesized virus.     

Three Epstein-Barr virus (EBV)-determined nuclear antigens induced by the BamHI E region of EBV DNA

In our previous study (Takada et al., 1986a), we showed that the BamHI E fragment of Epstein-Barr virus (EBV) DNA induces a nuclear antigen that is detected by human antisera against EBV-determined nuclear antigen (EBNA), when transfected into baby hamster kidney (BHK) cells. The present study shows that the sub-fragment containing the central open reading frame BERF2b of the BamHI E fragment (Baer et al., 1984) is responsible for nuclear antigen induction. In addition, 2 fragments corresponding to 2 other open reading frames of the BamHI E, BERFI and BERF4 also induce nuclear antigens upon transfection into BHK cells. These 3 antigens, designated RF2b, RFI and RF4 antigens, were serologically classified as EBNA and antigenically distinct. In immunoblotting analysis of latently EBV-infected BJ-B95-8 cells, 3 high-molecular-weight polypeptides (136, 142 and 147 kDa) were identified by anti-EBNA sera. Immunoblotting analysis of transfected BHK cells indicated that the RF2b antigen is 145 kDa in its native form and antigenically related to the 147-kDa protein of BJ-B95-8 cells. Although RFI and RF4 antigens were not detected by immunoblotting, reactivities of sera with RFI and RF4 antigens in the immunofluorescence test were correlated with those of sera with the 136- and 142-kDa polypeptides of BJ-B95-8 cells, respectively. The results suggest that 3 high-molecular-weight proteins of latently EBV-infected cells are encoded by 3 open reading frames of the BamHI E DNA fragment.