Evaluation of 12 commercial tests for detection of Epstein-Barr virus-specific and heterophile antibodies

Ten microbiological departments in Norway have participated in a multicenter evaluation of the following commercial tests for detection of Epstein-Barr virus (EBV)-specific and heterophile antibodies: CAPTIA Select viral capsid antigen (VCA)-M/G/EBNA (Centocor Inc.), Enzygnost anti-EBV/immunoglobulin M (IgM) and IgG (Dade Behring), Vironostika EBV VCA IgM/IgG/EBNA enzyme-linked immunosorbent assay (ELISA) (Organon Teknika), SEROFLUOR immunofluorescence assay and EBV Combi-Test (Institute Virion Ltd.), anti-EBV recombinant IgM- and IgG-early antigen/EBNA IgG ELISA (Biotest Diagnostics), EBV IgM/IgG/EBNA ELISA (Gull Laboratories), Paul-Bunnell-Davidsohn test (Sanofi Diagnostics Pasteur), Monosticon Dri-Dot (Organon Teknika), Avitex-IM (Omega Diagnostics Ltd.), Alexon Serascan infectious mononucleosis test (Alexon Biomedical Inc. ), Clearview IM (Unipath Ltd.), and Cards+/-OS Mono (Pacific Biotech, Inc.). The test panel included sera from patients with primary EBV infection, immunocompromised patients with recent cytomegalovirus infection, healthy persons (blood donors), and EBV-seronegative persons. Among the tests for EBV-specific antibodies the sensitivity was good, with only small differences between the different assays. However, there was a greater variation in specificity, which varied between 100% (Enzygnost) and 86% (Biotest). Tests for detection of heterophile antibodies based on purified or selected antigen (Avitex, Alexon, Clearview IM, and Cards+/-OS Mono) were more sensitive than the Paul-Bunnell-Davidsohn and Monosticon tests.     

Humoral immune response in Epstein-Barr virus infections. I. Elevated serum concentration of the IgG1 subclass in infectious mononucleosis and nasopharyngeal carcinoma

Using radial immunodiffusion we measured IgG subclass concentrations and studied their distribution in serum samples from patients with infectious mononucleosis (IM) and nasopharyngeal carcinoma (NPC), two Epstein-Barr virus (EBV)-associated diseases, in comparison with two control groups [completely anti-EBV negative persons and subjects carrying antibodies to the viral capsid antigen (VCA)]. Antibody titres to VCA and to the early antigen (EA) were determined by indirect immunofluorescence and revealed characteristic patterns for the respective diagnostic groups. Nephelometric assays served for quantitating total protein, albumin, total IgG, IgA and IgM in all the sera. In the IM and NPC groups the concentration of IgG1 was significantly elevated by more than 50% whereas the other three subclasses remained unchanged as compared with the controls. Correspondingly, we found a significant increase of total IgG in IM and NPC. In IM, the only disease where VCA-specific IgM antibodies have been reported to occur, IgM levels were markedly elevated. Our data suggest that the IgG1 subclass plays an important role in the humoral immune response to EBV-determined antigens and that it is possibly involved in the control of virus infection.     

Evaluation of 12 Commercial Tests for Detection of Epstein-Barr Virus-Specific and Heterophile Antibodies

Ten microbiological departments in Norway have participated in a multicenter evaluation of the following commercial tests for detection of Epstein-Barr virus (EBV)-specific and heterophile antibodies: CAPTIA Select viral capsid antigen (VCA)-M/G/EBNA (Centocor Inc.), Enzygnost anti-EBV/immunoglobulin M (IgM) and IgG (Dade Behring), Vironostika EBV VCA IgM/IgG/EBNA enzyme-linked immunosorbent assay (ELISA) (Organon Teknika), SEROFLUOR immunofluorescence assay and EBV Combi-Test (Institute Virion Ltd.), anti-EBV recombinant IgM- and IgG-early antigen/EBNA IgG ELISA (Biotest Diagnostics), EBV IgM/IgG/EBNA ELISA (Gull Laboratories), Paul-Bunnell-Davidsohn test (Sanofi Diagnostics Pasteur), Monosticon Dri-Dot (Organon Teknika), Avitex-IM (Omega Diagnostics Ltd.), Alexon Serascan infectious mononucleosis test (Alexon Biomedical Inc.), Clearview IM (Unipath Ltd.), and Cards±OS Mono (Pacific Biotech, Inc.). The test panel included sera from patients with primary EBV infection, immunocompromised patients with recent cytomegalovirus infection, healthy persons (blood donors), and EBV-seronegative persons. Among the tests for EBV-specific antibodies the sensitivity was good, with only small differences between the different assays. However, there was a greater variation in specificity, which varied between 100% (Enzygnost) and 86% (Biotest). Tests for detection of heterophile antibodies based on purified or selected antigen (Avitex, Alexon, Clearview IM, and Cards±OS Mono) were more sensitive than the Paul-Bunnell-Davidsohn and Monosticon tests.     

Performance of Two Commercially Available Automated Immunoassays for the Determination of Epstein-Barr Virus Serological Status

This study evaluated the performance of two automated Vidas (V) and Liaison (L) immunoassays for Epstein-Barr virus (EBV) serology. The detection of the viral capsid antigen (VCA) IgM, the VCA/early antigen (VCA/EA) IgG, and the Epstein-Barr nuclear antigen (EBNA) IgG was assessed on 526 sera collected for routine EBV testing in immunocompetent subjects. The determination of expected EBV status (186 EBV primary infections, 183 past EBV infections, and 157 EBV-seronegative individuals) was based on results of routine laboratory enzyme immunoassays (EIAs) together with clinical data. The sensitivity and specificity of each individual marker were determined in comparison to the expected EBV status. The agreement between the V and L profiles and the expected EBV status was established through the interpretation of combinations of the different EBV markers. Statistically significant differences between the two tests were found for the specificity of the VCA IgM marker (96.2% for V versus 93.2% for L), the sensitivity of the VCA/EA IgG marker (89% for V versus 94% for L), and the specificity of the EBNA IgG marker (96.5% for V versus 74.2% for L). The results determined for the two assays with respect to overall agreement with the established expected EBV status were not significantly different (89.7% for V versus 88.2% for L), with discrepancies mainly observed in sera referenced as primary infections. These findings demonstrated the similar performances of the Vidas and the Liaison assays for the establishment of an EBV serological status using the VCA, EA, and EBNA markers.     

Comparative evaluation of three commercial tests for detection of heterophile antibody in patients with infectious mononucleosis.

Infectious mononucleosis (IM) is caused by the Epstein-Barr virus. Commonly used laboratory tests used for diagnosis of IM include a screening test based on the observation that horse erythrocytes are agglutinated by the Paul-Bunnell antibody found in the serum of patients with IM. This study evaluated two latex agglutination (LA) kits for IM, Monolatex (Wampole Laboratories) and Immunoscan-IM (American MicroScan) (formerly Monogen; Biokit, S.A.), and compared them with Monospot (Ortho Diagnostic Systems) results on 220 patient sera. Discrepancies in the three test results were resolved with complete Epstein-Barr virus antibody profiles. They indicated that any of the three kits tested can be successfully used as a screening test for IM. The advantage of the LA kits is that no differential absorption step is necessary. When discrepancies were resolved, sensitivity and specificity of both LA kits were greater than 93%.     

Performance of the Architect EBV Antibody Panel for Determination of Epstein-Barr Virus Infection Stage in Immunocompetent Adolescents and Young Adults with Clinical Suspicion of Infectious Mononucleosis

The Architect EBV antibody panel is a new chemiluminescence immunoassay system used to determine the stage of Epstein-Barr virus (EBV) infection based on the detection of IgM and IgG antibodies to viral capsid antigen (VCA) and IgG antibodies against Epstein-Barr nuclear antigen 1 (EBNA-1). We evaluated its diagnostic accuracy in immunocompetent adolescents and young adults with clinical suspicion of infectious mononucleosis (IM) using the RecomLine EBV IgM and IgG immunoblots as the reference standard. In addition, the use of the antibody panel in a sequential testing algorithm based on initial EBNA-1 IgG analysis was assessed for cost-effectiveness. Finally, we investigated the degree of cross-reactivity of the VCA IgM marker during other primary viral infections that may present with an EBV IM-like picture. High sensitivity (98.3% [95% confidence interval {CI}, 90.7 to 99.7%]) and specificity (94.2% [95% CI, 87.9 to 97.8%]) were found after testing 162 precharacterized archived serum samples. There was perfect agreement between the use of the antibody panel in sequential and parallel testing algorithms, but substantial cost savings (23%) were obtained with the sequential strategy. A high rate of reactive VCA IgM results was found in primary cytomegalovirus (CMV) infections (60.7%). In summary, the Architect EBV antibody panel performs satisfactorily in the investigation of EBV IM in immunocompetent adolescents and young adults, and the application of an EBNA-1 IgG-based sequential testing algorithm is cost-effective in this diagnostic setting. Concomitant testing for CMV is strongly recommended to aid in the interpretation of EBV serological patterns.     

Serological diagnosis of Epstein-Barr virus infection by novel ELISAs based on recombinant capsid antigens p23 and p18

A new pair of Epstein-Barr virus ELISAs (Biotest Anti-EBV VCA IgG and VCA IgM ELISA) was evaluated for usefulness for routine diagnosis of acute EBV infections. The ELISAs are based on two viral capsid antigens (VCA), p23 (BLRF2, full-length) and p18 (BFRF3, carboxy-half), that are combined by autologous gene fusion. In total, 179 sera were tested in direct comparison with classical VCA immunofluorescence assays (IFA). With the help of clinical data and additional reference serology, i.e., heterophile antibodies, anti-EA IgG (IFA) and anti-EBNA-1 IgG (ELISA), the patients were divided into the following categories: seronegatives (46), acute primary infections (67), previous infections (39), suspected reactivations (20) and constellations with intermediate serological patterns (7). The VCA IgG and VCA IgM ELISAs showed overall agreement to IFA of 95.0% and 94.4%, respectively. The calculated analytical performance (sensitivity; specificity) of VCA IgG and VCA IgM was 94.0%; 97.8% and 97.1%; 96.5%, respectively. A certain delay in seroconversion of anti-p23-p18 IgG may account for a significant difference in sensitivity of the VCA IgG ELISA between primary (88.4%) and previous infections (100%). In summary, the new recombinant VCA ELISAs yielded good correlation to VCA IFA and in combination with EBNA-1 IgG allow rapid, sensitive, and specific diagnosis of infectious mononucleosis or EBV immune status in general.     

Evaluation of an Immunofiltration Assay That Detects Immunoglobulin M Antibodies against the ZEBRA Protein for the Diagnosis of Epstein-Barr Virus Infectious Mononucleosis in Immunocompetent Patients

The performance of an immunofiltration assay (IMFA) that detects immunoglobulin M (IgM) antibodies to the Epstein-Barr virus (EBV) ZEBRA (BamHI Z EBV replication activator) protein was evaluated for the diagnosis of EBV infectious mononucleosis (IM) in immunocompetent patients. The test panel consisted of 47 sera displaying an EBV-specific antibody profile compatible with an acute primary EBV infection from patients with clinical and biological features of EBV IM, 20 sera from healthy individuals either with a past EBV infection or who were EBV seronegative, 20 sera displaying an equivocal EBV antibody pattern (viral capsid antigen IgG positive [VCA IgG⁺], VCA IgM⁺, and EBV nuclear antigen-1 IgG⁺), and 15 sera obtained from patients with a mononucleosis-like syndrome owing to cytomegalovirus, human herpesvirus 6, or parvovirus B19. Overall, the sensitivity and the specificity of the assay were found to be 92.5%, and 97.3%, respectively. The sensitivity of the assay for the diagnosis of heterophile antibody-negative EBV IM was 86.2%. The IMFA is rapid, easy to perform, and, thus, suitable for point-of-care testing, and it may be used as a first-line test for the diagnosis of acute EBV IM in immunocompetent patients.Diagnosis of Epstein-Barr virus (EBV) infectious mononucleosis (IM) is commonly made on the basis of characteristic clinical manifestations and the detection of heterophile antibodies (HA). Nevertheless, HA may be absent, particularly in young children (14) but also in as many as 20% of adults with EBV IM (7). In these cases, demonstration of the presence of EBV viral capsid antigen (VCA) immunoglobulin G (IgG) and/or IgM antibodies, along with the absence of IgG antibodies to EBV nuclear antigen-1 (EBNA-1), allows the diagnosis of EBV primary infection (9). Detection of EBV-specific antibodies is accomplished by the use of commercial enzyme immunoassays, indirect immunofluorescence assays, line blot immunoassays (9), or, as established more recently, a multiplexed bead assay (3). These methods have long turnaround times, are labor-intensive, or require specific instruments or skilled technologists for their performance. In addition, interpretation of EBV VCA IgG/IgM and EBNA-1 IgG reactivity profiles is not always straightforward (9).The ZEBRA (BamHI Z EBV replication activator) protein is encoded by the immediate early BZLF1 gene. ZEBRA is expressed during the lytic cycle in EBV-permissive cells and plays a critical role in transactivating several immediate early, early, and late EBV genes (5). Antibodies against ZEBRA are produced during primary EBV infection (11, 15, 18), and thus, the detection of ZEBRA-specific IgMs may allow an early diagnosis of EBV IM. In the present study, we evaluated a rapid and easy-to-perform immunofiltration assay (IMFA) detecting IgMs to the EBV ZEBRA protein for the biological diagnosis of IM in immunocompetent patients.     

Comparative studies of three serologic methods for the measurement of Mycoplasma pneumoniae antibodies

Antibodies against Mycoplasma pneumoniae in the sera of patients and normal adult controls were measured by a standard complement fixation (CF) test, a commercial immunofluorescence (IF) test (CROWNTITRE), and a commercial enzyme-linked immunosorbent assay (ELISA) (MYCOPLASMELISA). The findings showed that, in the control sera, 269 of 277 (97%) had negative results for CF antibodies. Of the 320 controls tested by the IF assay, all (100%) had negative results for IgM antibodies and 314 (98%) had negative results for IgG antibodies. Only 6 of the 201 (3%) controls by the ELISA were classified as negative/equivocal. Among the 450 patient sera, 105 (23%) had positive results for CF antibodies, and 158 (35%) had positive results for IgG and/or IgM membrane antibodies by the IF test; 424 of these patients' sera were also tested by the ELISA, and 397 (94%) of them were found to have positive results for anti-M. pneumoniae IgG antibodies. If the CF test were chosen as the standard for comparison, the IF test would have a sensitivity of 87% and a specificity of 81% and the ELISA would have a sensitivity of 71% and a specificity of 80%, provided an adjustment in the threshold ELISA-positive value was made. A single positive M. pneumoniae membrane IgM antibody titer appeared to be valuable for a presumptive diagnosis of an ongoing infection; 41 of 47 (87%) of the IgM-positive results in the paired sera were supported by a fourfold increase or a stable high level of CF antibody titer.     

应用基于Logistic回归的ROC曲线评价鼻咽癌EB病毒抗体联合检测

目的 以基于Logistic回归的受试者工作特征(ROC)曲线分析方法,评价VCA/IgA、EA/IgA、Rta/IgG和EBNA1/IgA等EB病毒抗体不同组合在鼻咽癌诊断中的价值.方法 收集211例初治鼻咽癌和203例相似症状的非鼻咽癌病例的血清,采用免疫酶法检测VCA/IgA及EA/IgA,酶联免疫吸附法检测Rta/lgG和EBNA1/IgA.对各种的抗体组合建立Logistic回归模型,以预测概率为分析指标,应用ROC曲线分析,评价不同组合对鼻咽癌的诊断价值.结果 单一指标评价,VCA/IgA敏感度最高(98.1%),EA/IgA特异度最高(98.5%).以基于Logistic同归的ROC曲线分析各项组合,敏感度和特异度均有所提高.双指标组合中,VCA/IgA+Rta/lgG组合的ROC曲线下面积(AUC)为0.991,诊断效能最高,敏感度、特异度及约登指数分别为94.8%、98.0%及0.928.VCA/IgA+Rta/IgG+EBNA1/IgA组合和4项指标组合的敏感度、特异度及约登指数分别为94.8%、98.5%、0.933和96.7%、97.0%、0.937;这两种多指标组合的AUC与VCA/IgA+Rta/IgG组合比较差异均无统计学意义(P＞0.05).结论 基于Logistic回归的ROC曲线分析方法可以为多指标联合诊断试验提供更客观的综合分析,VCA/IgA和Rta/IgG联合检测具有互补作用,是鼻咽癌血清学诊断的合适组合.     

Measurement of EBV-IgG anti-VCA avidity aids the early and reliable diagnosis of primary EBV infection

Current serological methods for the diagnosis of Epstein-Barr virus (EBV) infection still differentiate poorly between primary infection and reactivation. This is particularly true when IgG and IgM antibodies are present simultaneously and only a single serum sample is provided for analysis. The demonstration of the IgG avidity state has the potential to distinguish recent from past or reactivated infection. An analysis of the kinetics of avidity maturation of anti-VCA antibodies in primary EBV infection was undertaken with longitudinally collected sets of sera from 28 well-characterised EBV cases and in sera from 35 cases with previous EBV infection and recent primary infection due to HIV, CMV, or hepatitis A. Antibodies directed against the viral capsid antigen (VCA) and Epstein-Barr nuclear antigen (EBNA-1) were sought, using a commercial enzyme immunoassay (EIA). In parallel with standard IgG anti-VCA detection, serum was incubated with 8 M urea to disrupt low-avidity complexes to allow calculation of the percentage avidity. In cases with primary EBV infection, the mean avidity rose from 54% at 6 weeks to 82% by 28 weeks after the onset of symptoms, but remained lower than that of the control sera (96%). The addition of the avidity measurement improved the sensitivity of IgG and IgM anti-VCA testing in diagnosis of primary EBV infection from 93% to 100%. The specificity of IgM anti-VCA testing alone was poor, with 14 of 35 cases (49%) demonstrating false-positive results, but it improved to 97% by the demonstration of high-avidity IgG anti-VCA. The combination of negative IgG anti-EBNA and low-avidity IgG anti-VCA had a sensitivity and specificity of 100%. The routine addition of IgG anti-VCA avidity estimation to diagnostic EBV serology is recommended.     

Serodiagnosis of Epstein-Barr Virus Infection by Using Recombinant Viral Capsid Antigen Fragments and Autologous Gene Fusion

Using recombinant 15- to 30-kDa fragments and fusion with glutathione S-transferase (GST), we investigated the seroreactivity of three large structural proteins of Epstein-Barr virus (EBV), p150 (BcLF1, capsid), p143 (BNRF1, tegument), and gp125 (BALF4, membrane) in Western blots. None of 13 fragments tested, however, was qualified for diagnostic application. In contrast, the two small viral capsid antigens (VCA), p18 (BFRF3) and p23 (BLRF2), demonstrated sensitive (100%) EBV-specific immunoglobulin G (IgG) reactivities. While p18 additionally showed maximum sensitivity for IgM detection, the IgM sensitivity of p23 was restricted (44%). An autologous fusion protein, p23-p18, which consists N-terminally of full-length p23, followed by the carboxy half of p18, was constructed. This antigen was subjected to indirect VCA enzyme-linked immunosorbent assays (ELISAs), for IgG and IgM, as well as to a micro-capture (microc) IgM ELISA. All assays were found to be 100% specific when EBV-negative sera were tested. Using sera from previously infected individuals, the p23-p18 fusion revealed an improved IgG sensitivity of 99% compared to sensitivities of 97 and 93% for the single antigens p18 and p23, respectively. The sensitivity and specificity of the indirect IgM ELISA with samples of primary and past infections, respectively, were 100%. The microc principle for IgM overcame completely the interference by rheumatoid factors. Compared to the specificity of the indirect IgM version, the specificity with sera collected from rheumatoid arthritis patients increased from 48 to 100%. In summary, the p23-p18 IgG and microc IgM ELISAs showed excellent performances and are promising new diagnostic tests for the detection of EBV-specific antiviral capsid antibodies.     

Detection of infectious mononucleosis heterophil antibody by a rapid, standardized enzyme-linked immunosorbent assay procedure

A rapid, specific, sensitive, standardized, a reproducible enzyme-linked immunosorbent assay (ELISA) procedure has been developed for detecting the heterophil antibody associated with infectious mononucleosis (IM). The IM heterophil antibody used for the solid phase was purified from bovine erythrocyte stroma. The test uses heavy-chain-specific anti-immunoglobulin M (IgM) labeled with alkaline phosphatase and three 10-min incubations. The quantitative results correlated well with horse erythrocyte agglutination titers. Absorption tests confirmed the specificity of the ELISA reactions for IM heterophil antibodies. Neither very high levels of IgM in myeloma sera nor high levels of rheumatoid factor caused false-positive reactions. A number of probable IM cases were encountered which positive by ELISA but negative by the horse erythrocyte slide agglutination test. Absorption studies indicated that these were true-positives for the IM heterophil antibody. The IM heterophil antibodies were confirmed to be predominantly of the IgM class, but moderate proportions of the IgM class were sometimes encountered.     

Evaluation of a dipstick ELISA and a rapid immunochromatographic test for diagnosis of Dengue virus infection

Here we report standardization of a dipstick enzyme-linked immunosorbent assay (ELISA, Dipstick ELISA) and its comparative evaluation with a commercial Rapid PanBio Immunochromatographic test (IC test) for detection of Dengue (DEN) virus-specific IgM and IgG antibodies in patient sera. Among crude and purified viral antigens prepared from mouse brains or cell cultures, a DEN virus type 2 antigen purified from cell cultures by sucrose density gradient centrifugation was found superior in terms of the signal/ noise (S/N) ratio in the assay system. The sensitivity of detection of the virus by specific IgM antibody was improved by removal of IgG from patient sera prior to testing. The evaluation of the Dipstick ELISA by use of 156 serum samples revealed an overall accordance of 96% and 93% with the IC test in detection of IgM antibodies to DEN viruses (IgM antibodies) and IgG antibodies to DEN viruses (IgG antibodies), respectively. The sensitivity of the Dipstick ELISA and the IC test with reference to the mu-capture ELISA was 83% and 87%, respectively, with a specificity of 98% in both cases. The sensitivity of the Dipstick ELISA with reference to the IC test in detecting IgM and IgG antibodies was 84% and 94%, respectively, and the specificity of the Dipstick ELISA was 98% and 92%, respectively.     

Measurement of antibodies to avian influenza virus A(H7N7) in humans by hemagglutination inhibition test

During the epizootic of highly pathogenic avian influenza A(H7N7) in 2003 in The Netherlands, RT-PCR and culture confirmed infection was detected in 89 persons who were ill. A modified hemagglutination inhibition (HI) test using horse erythrocytes and 2 hemagglutinating units of virus was applied to assess retrospectively the extent of human (subclinical) infection. Validation of the HI-test with sera from 34 RT-PCR and culture confirmed A(H7) infected persons and sera from 100 persons from a human influenza vaccine trial in autumn 2002 showed that this HI-test had a sensitivity of 85% and a specificity of 100% when using a cut-off titer of > or =10. Using this cut-off value, A(H7) specific antibodies were detected in 49% of 508 persons exposed to poultry and in 64% of 63 persons exposed to A(H7) infected persons. Correlation of seropositivity with the occurrence of eye symptoms in exposed persons who had not received antiviral prophylaxis and of reduced seropositivity with taking antiviral prophylaxis provided further evidence that the A(H7) HI antibody titers were real. In conclusion, by applying an HI-test using horse erythrocytes human antibodies against the avian A(H7N7) virus were detected with high sensitivity and specificity in an unexpectedly high proportion of exposed persons.     

Studies on the Paul-Bunnell antigen-antibody system. III. Detection of a Paul-Bunnell-related antigen in syphilis and leprosy

By means of double-diffusion precipitation tests in agarose gel with selected infectious mononucleosis (IM) sera, an antigen was detected in sera of patients with syphilis and leprosy. Of 378 syphilis sera tested 39 (10.3%) and of 36 leprosy sera 7 (19.4%) gave positive results. Sera of patients with various other diseases were negative. This antigen was demonstrable in sera of patients with both early and late syphilis and was shown to be unrelated to cardiolipin. Absorption studies of the IM sera with bovine and sheep erythrocytes and guinea pig tissues revealed that this antigen was identical with or closely related to the previously described B antigen of the Paul-Bunnel (P-B) antigenic complex and distinct from heterophile antigens of Hanganutziu-Deicher (H-D) and of Forssman specificity. However, 2 of 89 syphilis sera without the P-B antigen were shown to contain antigen(s) of H-D specificity. None of the syphilis or the leprosy sera contained P-B antibodies, but H-D antibodies were found in 13% of syphilis sera.     

The use of semi-automated EBV IgG avidity determination for the diagnosis of infectious mononucleosis

The diagnosis of acute Epstein-Barr virus (EBV) infection is based frequently on the combination of positive viral capsid antigen (VCA) IgM antibodies and negative EB viral nuclear antigen 1 (EBNA-1) IgG antibodies. However, both VCA IgM and EBNA-1 IgG can provide false positive and false negative results. Therefore, situations in which the EBV serology remains unclear are not uncommon. Determination of EBV IgG avidity can clarify the EBV status in these patients. So far, mainly immunofluorescence assays have been used for this purpose. These tests are laborious, their evaluation is subjective, and automation is difficult. Therefore, two commercially available microtiter plate enzyme immunoassays (EIA) were compared for their usefulness for semi-automated EBV IgG avidity determination. One assay is based on a mixture of EBV antigens, the other assay uses a synthetic peptide of the VCA-complex. Patient sera of confirmed acute and past EBV infections were tested for avidity by both assays. The results with the antigen mixture assay proved to be highly sensitive (100%) and specific (100%). Avidity index calculations on the basis of one-point-quantification titers gave better results than calculations using OD values. Determination of EBV IgG avidity by the peptide assay was complicated by the fact that it was less sensitive than the antigen mixture assay for IgG detection in acute EBV infections. On the other hand, about 30% of the samples had to be retested with the peptide assay in a higher dilution because the IgG units in initial testing fell outside the range covered by the standard curve. Using OD values of the peptide EIA, the sensitivity was 99% but the specificity of detection of acute EBV infections was only 86%. Thus, while the peptide EBV avidity assay is unsuitable as a confirmatory assay, avidity testing with the antigen mixture assay is a useful tool to resolve equivocal EBV serologies. Avidity assays on the basis of EIA can be automated which should lead to wider use of this methodology. J. Med. Virol. 54:145–153, 1998. © 1998 Wiley-Liss,Inc.     

Heterophile IgM, IgA, and IgE antibodies in infectious mononucleosis

Fifty infectious mononucleosis (IM) and 150 non-infectious mononucleosis (non-IM) sera were tested by a hemadsorption immunocapture test (HIT) for the detection of heterophile antibodies of IgM, IgA, and IgE classes. The specificity of Paul-Bunnell (PB) antibodies was ascertained by a differential absorption test. IgM PB-antibodies were demonstrated in 100% of IM sera, IgA in 92%, and IgE in 88%. PB antibodies were present only in IM sera; but other heterophile antibodies of IgM (68%), IgA (6.7%), and IgE (9.3%) classes were demonstrated in non-IM sera. IgA and IgE heterophile antibodies were thus present, especially in IM sera. HIT was found to be more sensitive than Paul-Bunnell Davidsohn test (PBD test) and suitable for the diagnosis of infectious mononucleosis.     

Epstein-Barr virus serology in the diagnosis of nasopharyngeal carcinoma

The antibody levels to viral capsid antigen (VCA) and early antigen (EA) of Epstein-Barr virus (EBV) in 164 nasopharyngeal carcinoma (NPC) patients from Sarawak, East Malaysia were significantly higher than those in 147 sex, age and ethnically matched healthy controls. As diagnostic markers of NPC, IgG/VCA at reciprocal titers > or =160 was the most sensitive (89%, with 98% specificity), while IgA/EA at > or =5 was the most specific (100%) but the least sensitive (75%). The sensitivity and specificity of IgA/VCA at reciprocal titers > or =10 were 84% and 97%. IgA/VCA has an advantage over IgG/VCA despite the slightly lower sensitivity due to its consistently more distinct fluorescence reaction. The sensitivity and specificity can be marginally improved by a combination of two tests.     

Reticulate bodies as single antigen in Chlamydia trachomatis serology with microimmunofluorescence.

Formalin-fixed, purified reticulate bodies (RB) of Chlamydia trachomatis immunotype C/TW-3/OT were used as a serological test antigen in the microimmunofluorescence test. The sensitivity and specificity of the RB antigen were compared to elementary bodies (EB) used as antigens in the detection of C. trachomatis antibodies in human sera by microimmunofluorescence. RB reacted with all known C. trachomatis immunotypes with the same sensitivity as the homotypic EB. In routine serology with sera and endocervical secretions, the RB antigen had a sensitivity similar to that of the EB in detecting serum antibodies, endocervical secretion antibodies, and antibodies of immunoglobulin M and G classes. No false-positive reactions were detected with control sera. All positive reactions showed type-specific antibodies against an EB immunotype. RB seemed to demonstrate chlamydial group reactivity; sera from 10 psittacosis patients diagnosed clinically and serologically by complement fixation showed five positive, three equivocal, and two negative reactions. By immunofluorescence, RB appeared as distinct rings demonstrating uniform peripheral surface fluorescence at their rims. The EB appeared as pinpoint-sized dots. C/TW-3/OT RB used as a single test anitgen should provide a simple and sensitive serological assay for the detection of C. trachomatis antibody.