Rapid Detection of Antigenic Diversity of Akabane Virus Isolates by Dot Immunobinding Assay Using Neutralizing Monoclonal Antibodies

Akabane (AKA) virus is an arthropod-borne virus belonging to the Simbu group of the genus Bunyavirus. Neutralizing monoclonal antibodies (MAbs) against AKA virus were prepared, and the neutralizing epitopes of the virus were defined by competitive binding assay. Five distinct antigenic domains were identified and were designated A, B, C, D, and E. Domains A and C consisted of two epitopes each. It was demonstrated that seven neutralizing epitopes exist on the G1 glycoprotein of AKA virus. Dot immunobinding assays (DIAs) were performed with MAbs which recognize these seven neutralizing epitopes. The results were similar to those obtained by enzyme-linked immunosorbent assay. DIAs were performed using two Australian strains, one isolate from Taiwan, and isolates from Japan collected between the years 1959 and 1994, for a total of 63 isolates. The MAb response patterns were divided into five groups: the OBE-1 strain, the JaGAr39 strain, the Iriki strain, a group which consisted of features between those of the JaGAr39 strain and Iriki strain groups, and a group which did not belong to any of these patterns. The isolates which showed patterns similar to that of the JaGAr39 strain were found mostly among the isolates collected in 1974 and 1990, and isolates with patterns of MAb responses similar to the pattern of the Iriki strain were found mostly in the 1985 isolates. Those showing patterns in between were found mostly around 1977, 1987, and 1994. The results show that DIA can be used to effectively compare the antigenicities of AKA virus isolates within a few hours, even with lesser amounts of virus culture than is required for other assays.     

用碱性磷酸酶免疫斑点试验检测小鼠血清中的HFRS病毒抗体

将ＨＦＲＳ病毒陈株可溶性抗原吸附于硝酸纤维素膜上，建立了用碱性磷酸酶免疫斑点试验（ＡＰ－ＤＩＢＡ）检测小鼠血清中的ＨＦＲＳ病毒抗体的方法，并与应用辣根过氧化物酶的免疫斑点试验（ＨＲＰ－ＤＩＢＡ）作了比较。结果表明，ＡＰ－ＤＩＢＡ具有特异性；其敏感性高于ＨＲＰ－ＤＩＢＡ。该法可用于不同ＨＦＲＳ病毒株感染小鼠血清中抗ＨＦＲＳ病毒抗体的检测。     

Transmission of Human T-Cell Lymphotropic Virus Type 1 Tax to Rabbits by tax-Only-Positive Human Cells

The human T-cell lymphrotropic virus type 1 (HTLV-1) is causally related to adult T-cell leukemia and lymphoma and the neurodegenerative diseases tropical spastic paraparesis and HTLV-1-associated myelopathy. In the United States the prevalence of infection has been estimated to range from 0.016 to 0.1% on the basis of serologic tests for antibodies to the viral structural proteins. Blood from donors positive for antibodies to HTLV-1 or HTLV-2 is not used for transfusion. However, patients with the cutaneous T-cell lymphoma mycosis fungoides (MF) are HTLV-1 and -2 seronegative yet harbor proviral sequences identical to those that encode the HTLV-1 transactivating and transforming gene product p40tax in their peripheral blood mononuclear cells (PBMCs), and they usually have antibodies to p40^tax. Moreover, a study of 250 randomly selected blood donors revealed that approximately 8% of these seronegative individuals also had HTLV-1 tax sequences and antibodies to p40^tax, while they lacked sequences and antibodies related to gag, pol, or env. Thus, it seemed important to determine whether the “tax-only” state can be transmitted by transfusion. To this end, PBMCs from HTLV-1 and -2 seronegative tax-only-positive MF patients or from healthy tax-only-positive blood donors were injected into adult rabbits, an established animal model for HTLV-1 infection. The PBMCs of all injected rabbits became tax sequence positive. These observations suggest that HTLV-1 tax can be transmitted by tax-only-positive mononuclear cells.     

滴金免疫渗滤法检测广州管圆线虫抗体的实验研究

目的建立一种检测广州管圆线虫抗体的简便、快速、敏感、特异的新方法。方法以广州管圆线虫成虫抗原为包被抗原，金标记SPA为显色剂，建立检测广州管圆线虫抗体的滴金免疫渗滤法（DIGFA）；并以ELISA平行检测比较。结果用DIGFA检测广州管圆线虫实验感染鼠血清50份，其阳性检出率为96．0％，50份正常鼠血清的阴性符合率达100％。DIGFA分别检测蛲虫阳性鼠血清13份。绦虫阳性鼠血清11份和粪类圆线虫阳性鼠血清18份，前二者均为阴性，后者交叉反应率为5．6％；DIGFA与ELISA平行检测30份广州管圆线虫阳性鼠血清，两法符合率达96，7％。结论DIGFA与ELISA有相似的敏感性和特异性。且具有简便、快速、不需特殊仪器设备等优点。是一种检测广州管圆线虫抗体的新方法。     

Human T-Cell Lymphotropic Virus Type 1 Tax among American Blood Donors

In the United States, all blood used for transfusion is tested for the presence of antibodies to the structural components of the human T-cell lymphotropic viruses types 1 and 2 (HTLV-1 and -2). Based on such serologic tests, the prevalence of HTLV-1 infection is estimated to range from 0.016 to 0.1%. As a consequence of studies of patients with mycosis fungoides and some of their healthy relatives who are antibody negative but were found to carry the tax sequence of HTLV-1 in their lymphocytes and who had antibodies to the p40^tax protein, a study was undertaken to determine the prevalence of the “tax-only” state in 250 healthy blood donors and other volunteers. Using PCR and Southern analysis for cell lysates and using Western blotting for plasmas, 8.6% of the blood donors proved to be tax sequence positive and antibody positive. Sequence analysis of specimens from 22 individuals proved that 20 of the sequences were homologous with that of HTLV-1 while 2 resembled the HTLV-2 sequence. The latter were obtained from volunteers of Indian origin. The possible clinical significance of the tax-only carrier state is discussed.     

Heterophile Antibodies to Bovine and Caprine Proteins Causing False-Positive Human Immunodeficiency Virus Type 1 and Other Enzyme-Linked Immunosorbent Assay Results

Heterophile antibodies are a well-recognized cause of erroneous results in immunoassays. We describe here a 22-month-old child with heterophile antibodies reactive with bovine serum albumin and caprine proteins causing false-positive results to human immunodeficiency virus type 1 and other infectious serology testing.     

Immune Response against the Exp-1 Protein of Plasmodium falciparum Results in Antibodies That Cross-React with Human T-Cell Lymphotropic Virus Type 1 Proteins

To examine the role of the Plasmodium falciparum Exp-1 blood-stage protein in producing antibodies that cross-react with human T-cell lymphotropic virus type I (HTLV-I) proteins, we studied sera from Indonesian volunteers who seroconverted to malaria after transmigrating to an area where malaria is hyperendemic. Samples from Philippine volunteers, that were used in a prior study that examined malaria antibodies that cross-react with HTLV-I proteins, were also used. Eighty-three percent of the Indonesian transmigrants developed antibodies against the malaria Exp-1 protein by 6 months postmigration. Of these malaria seroconverters, 27% developed false-positive HTLV-I enzyme immunoassay (EIA) immunoreactivity, as indicated by indeterminate HTLV-I Western blot banding patterns. Five of the six Philippine samples tested were HTLV-I EIA false positive and Western blot indeterminate. When a recombinant Exp-1 protein was used in blocking experiments, the HTLV-I Western blot immunoreactivity of sera from both groups was either completely eliminated or greatly reduced. No effect on the Western blot immunoreactivity of truly HTLV-I-positive sera was seen. To determine if immunization with the recombinant Exp-1 protein could elicit the production of HTLV-I antibodies, six mice were inoculated with the recombinant protein. Following administration of three 50-μg doses of the protein, four of the six mice developed antibodies that cross-reacted with HTLV-I proteins on Western blot. These results indicate that the immune response against the malaria Exp-1 protein may result in HTLV-I-cross-reacting antibodies that can lead to false-positive EIA and indeterminant Western blotting results.     

Flexible Label-Free Quantitative Assay for Antibodies to Influenza Virus Hemagglutinins

During the initial pandemic influenza H1N1 virus outbreak, assays such as hemagglutination inhibition and microneutralization provided important information on the relative protection afforded by the population''s cross-reactivity from prior infections and immunizations with seasonal vaccines. However, these assays continue to be limited in that they are difficult to automate for high throughput, such as in pandemic situations, as well as to standardize between labs. Thus, new technologies are being sought to improve standardization, reliability, and throughput by using chemically defined reagents rather than whole cells and virions. We now report the use of a cell-free and label-free flu antibody biosensor assay (f-AbBA) for influenza research and diagnostics that utilizes recombinant hemagglutinin (HA) in conjunction with label-free biolayer interferometry technology to measure biomolecular interactions between the HA and specific anti-HA antibodies or sialylated ligands. We evaluated f-AbBA to determine anti-HA antibody binding activity in serum or plasma to assess vaccine-induced humoral responses. This assay can reveal the impact of antigenic difference on antibody binding to HA and also measure binding to different subtypes of HA. We also show that the biosensor assay can measure the ability of HA to bind a model sialylated receptor-like ligand. f-AbBA could be used in global surveillance laboratories since preliminary tests on desiccated HA probes showed no loss of activity after >2 months in storage at room temperature, indicating that the same reagent lots could be used in different laboratories to minimize interlaboratory assay fluctuation. Future development of such reagents and similar technologies may offer a robust platform for future influenza surveillance activities.Vaccination, the cornerstone of public health intervention, helps prevent influenza morbidity and mortality. Effective vaccines induce protective immunity which is correlated with the presence of virus-specific antibodies (Abs) in serum that are directed against the external coat proteins of the virion, hemagglutinin (HA) and, to a lesser extent, neuraminidase (NA). HA is the principal antigen on the viral surface, and neutralizing antibodies are usually directed to hypervariable epitopes located in or near the HA receptor-binding site (RBS) and act to prevent host infection by blocking virus binding to the host cell (17). For this reason, induction of HA-specific antibodies that interfere with virus entry is used as a correlate of vaccine protective efficacy.Influenza viruses are characterized by their rapid antigenic change as a result of their high mutation frequency. Therefore, the composition of influenza virus vaccines requires frequent updates (every 2 years on average for H3N2) to match their antigenicity as closely as possible to that of the variant viruses most prevalent in the population (25). Analysis of antibody responses that correlate with protective immunity to influenza virus vaccination is an important element in the assessment of the potential impact of viral antigenic drift. The hemagglutination inhibition (HI) test is the most widely used serological test for the detection of anti-influenza virus antibodies (13, 36) and is used routinely to determine the serological outcome of vaccinations. The assay itself is technically simple but difficult to automate and standardize. In addition, interpretation of results can be affected by virus passage, antibody source (species), and variability between red blood cells from different species (19, 36).The continuing but sporadic human infections with H5N1 avian influenza viruses reported since 1997 have revealed that HI assays are less sensitive in detecting antibodies against avian influenza viruses (2, 16, 23, 24, 28) than are alternative assays such as the microneutralization (MN) test (24). The MN test, however, is technically very demanding and is currently performed only as a reference test on a small number of serum specimens. As virus-neutralizing activity of antiserum is mediated in part by blocking virus-receptor interactions, the results of the HI test often correlate well with those of the MN test. Recent international laboratory network studies showed large intra- and interlaboratory assay variations for both the HI and MN assays (26), although for H5N1, such variability can be reduced through the availability of an antibody standard (27). Despite limited reproducibility between labs, the HI and MN tests still provide the best available data to inform global surveillance and aid in decisions to update the seasonal influenza virus vaccine composition. Indeed, their use in recent months has been critical in assessing immunity to the pandemic H1N1 virus in the human population (4, 20). It is clear that new technologies and assays are urgently needed to improve sensitivity, accuracy, and sample throughput, as well as reproducibility between labs. In addition, assays need to be flexible and adaptable to be able to analyze antibody responses to emerging viruses from sources other than avian populations (12).Here, we report on a flexible label-free and cell-free assay to determine the relative functional avidity of polyclonal serum antibodies binding to the major virion coat protein, HA. The flu antibody biosensor assay (f-AbBA) presented here utilizes an established recombinant baculovirus expression system for producing HA (32) in conjunction with label-free biolayer interferometry (BLI) technology from Fortebio Inc., an optical technique that analyzes the interference pattern of white light reflected from a layer of immobilized protein on the tip of a fiber optic biosensor (1, 7). Macromolecules binding to the biosensor tip produce an increase in optical thickness at the biosensor tip, which results in a wavelength shift (measured in nanometers) that can be followed in real time, allowing one to determine binding kinetics. BLI technology minimizes interference from extraneous materials present in solution. Only molecules that bind to or dissociate from the biosensor surface produce a signal change. Sample preparation time is reduced because crude mixtures such as cell lysates, patient serum, or hybridoma supernatants can be assayed. The platform is designed to simultaneously analyze eight samples as a dip-and-read format using commercially available 96-well plates. We have used it to measure the biomolecular interactions of recombinant HA with reactive antibodies from patient sera in real time, without the need for secondary reagents for detection. Results have good correlation with existing serological methods but with improved sensitivity and reproducibility. By comparing binding to different HAs, we can determine the polyclonal specificity, while limited studies with human sera from prior vaccination studies reveal a linear correlation between binding observed with the biosensor and corresponding HI titers as well as some cross-reactivity with the 2009 pandemic H1N1 virus HA. Initial stability tests suggest that reagents may be prepared and desiccated on probes and stored at ambient temperature for extended periods of time without loss of function, offering different laboratories the chance to utilize the same reagents and thus improve interlaboratory assay variation.     

Development and Evaluation of a Blocking Enzyme-Linked Immunosorbent Assay and Virus Neutralization Assay To Detect Antibodies to Viral Hemorrhagic Septicemia Virus

Viral hemorrhagic septicemia virus (VHSV) is a target of surveillance by many state and federal agencies in the United States. Currently, the detection of VHSV relies on virus isolation, which is lethal to fish and indicates only the current infection status. A serological method is required to ascertain prior exposure. Here, we report two serologic tests for VHSV that are nonlethal, rapid, and species independent, a virus neutralization (VN) assay and a blocking enzyme-linked immunosorbent assay (ELISA). The results show that the VN assay had a specificity of 100% and sensitivity of 42.9%; the anti-nucleocapsid-blocking ELISA detected nonneutralizing VHSV antibodies at a specificity of 88.2% and a sensitivity of 96.4%. The VN assay and ELISA are valuable tools for assessing exposure to VHSV.     

Development of a Luciferase Immunoprecipitation System Assay To Detect IgG Antibodies against Human Respiratory Syncytial Virus Nucleoprotein

The nucleoprotein of respiratory syncytial virus (RSV-N) is immunogenic and elicits an IgG response following infection. The RSV-N gene was cloned into a mammalian expression vector, pREN2, and the expressed luciferase-tagged protein (Ruc-N) detected anti-RSV-N-specific IgG antibodies using a high-throughput immunoprecipitation method (the luciferase immunoprecipitation system [LIPS]-N_RSV assay). The specificity of the assay was evaluated using monoclonal antibodies (MAbs) and monospecific pre- and postimmunization rabbit antisera. Blood serum samples from chimpanzees and humans with proven/probable RSV infection were also tested. The pre- and postimmunization serum samples from rabbits given human metapneumovirus (HMPV) or measles virus were negative when tested by the LIPS-N_RSV assay, while antisera obtained after immunization with either the RSV-A or RSV-B strain gave positive signals in a dose-dependent manner. RSV-N MAb 858-3 gave a positive signal in the LIPS-N_RSV assay, while MAbs against other paramyxovirus nucleoproteins or RSV-F or RSV-G did not. Serum samples from chimpanzees simultaneously immunized with vaccinia-RSV-F and vaccinia-RSV-G recombinant viruses were negative in the LIPS-N_RSV assay; however, anti-RSV-N IgG responses were detected following subsequent RSV challenge. Seven of the 12 infants who were seronegative at 9 months of age had detectable anti-RSV-N antibodies when they were retested at 15 to 18 months of age. The LIPS-N_RSV assay detects specific anti-RSV-N IgG responses that may be used as a biomarker of RSV infection.     

Detection of Serum Antibodies to Ovine Progressive Pneumonia Virus in Sheep by Using a Caprine Arthritis-Encephalitis Virus Competitive-Inhibition Enzyme-Linked Immunosorbent Assay

A competitive-inhibition enzyme-linked immunosorbent assay (cELISA) for detection of antibodies to the surface envelope (SU) of caprine arthritis-encephalitis virus (CAEV) was recently reported (L. M. Herrmann, W. P. Cheevers, T. C. McGuire, D. Scott Adams, M. M. Hutton, W. G. Gavin, and D. P. Knowles, Clin. Diagn. Lab. Immunol. 10:267-271, 2003). The cELISA utilizes CAEV-63 SU captured on microtiter plates using the monoclonal antibody (MAb) F7-299 and measures competitive displacement of binding of the anti-CAEV MAb GPB 74A by goat serum. The present study evaluated the CAEV cELISA for detection of antibodies to ovine progressive pneumonia virus (OPPV) in sheep. Three hundred thirty-two sera were randomly selected from 21,373 sheep sera collected throughout the United States to determine the sensitivity and specificity of cELISA and agar gel immunodiffusion (AGID) based on immunoprecipitation (IP) of [³⁵S]methionine-labeled OPPV antigens as a standard of comparison. A positive cELISA test was defined as >20.9 percent inhibition (% I) of MAb 74A binding based on two standard deviations above the mean % I of 191 IP-negative sheep sera. At this cutoff, there were 2 of 141 false-negative sera (98.6% sensitivity) and 6 of 191 false-positive sera (96.9% specificity). Sensitivity and specificity values for IP-monitored AGID were comparable to those for cELISA for 314 of 332 sera with unambiguous AGID results. Concordant results by cELISA and IP resolved 16 of the 18 sera that were indeterminate by AGID. Additional studies evaluated cELISA by using 539 sera from a single OPPV-positive flock. Based on IP of 36 of these sera, there was one false-negative by cELISA among 21 IP-positive sera (95.5% sensitivity) and 0 of 15 false-positives (100% specificity). We conclude that the CAEV cELISA can be applied to detection of OPPV antibodies in sheep with high sensitivity and specificity.     

Ultrastructural Pathology of Human T-Cell Lymphotropic Virus Type I Encephalomyelopathy in a White Patient with Adult T-Cell Leukemia/Lymphoma

A white patient with human T-cell lymphotropic virus type I (HTLV-I)-associated adult T-cell leukemia/lymphoma for 10 years died 4 months after the onset of spastic myelopathy. Ultrastructurally, the neuropathologic findings consisted of dystrophic neurites (spheroids) filled with neurofilaments or electron-dense bodies, intense astrocytic reaction with abundant formation of corpora amylacea, and multilamellar bodies. Demyelination and spongiform change were absent. Viruslike particles resembling HTLV-I were detected adjacent to brain endothelial cells.     

Ultrastructural Pathology of Human T-Cell Lymphotropic Virus Type I Encephalomyelopathy in a White Patient with Adult T-Cell Leukemia/Lymphoma

A white patient with human T-cell lymphotropic virus type I (HTLV-I)-associated adult T-cell leukemia/lymphoma for 10 years died 4 months after the onset of spastic myelopathy. Ultrastructurally, the neuropathologic findings consisted of dystrophic neurites (spheroids) filled with neurofilaments or electron-dense bodies, intense astrocytic reaction with abundant formation of corpora amylacea, and multilamellar bodies. Demyelination and spongiform change were absent. Viruslike particles resembling HTLV-I were detected adjacent to brain endothelial cells.     

Early Detection of Human Immunodeficiency Virus Type 1-Specific B-Lymphocyte-Derived Antibodies in a High-Risk Population

Diagnosis of acute human immunodeficiency virus (HIV) infection, a key driver of the HIV epidemic, remains a public health challenge. The PlasmAcute technology offers an opportunity to detect early anti-HIV antibody responses. B lymphocytes (B cells) were isolated from the blood of seronegative miners in South Africa by using the PlasmAcute method. B-cell lysates and paired sera were tested for anti-HIV-1 antibodies by two different enzyme-linked immunosorbent assays; immunoreactivity was confirmed by Western blotting. All volunteers were tested for HIV type 1 (HIV-1) viral load, p24 antigen, and CD4 count. Sera from HIV-seronegative men who had positive viral loads and were positive for p24 antigen were retested for anti-HIV antibodies after immune complex dissociation. Anti-HIV antibodies were detected in lysates from 16/259 subjects without immunoreactivity in paired sera. Four subjects, one of whom had a positive viral load initially, subsequently seroconverted. Six subjects showed transient anti-HIV-1 antibodies in the lysates and tested negative for all markers at the follow-up. Five subjects without follow-up data initially had lysate-positive/serum-negative samples, and these cases were classified as inconclusive. One subject had lysate antibodies and a detectable viral load but was seronegative at follow-up. In conclusion, lysate-derived anti-HIV-1 B-cell antibodies can be detected prior to seroconversion and earlier than or contemporary with HIV-1 RNA detection.According to the 2006 UNAIDS report on the global AIDS epidemic, an estimated 38.6 million people worldwide were living with human immunodeficiency virus (HIV) at the end of 2005 (31). It was also estimated that 4.1 million became newly infected with HIV and that 2.8 million lost their lives to AIDS in that year. Approximately 10% of the world''s population lives in sub-Saharan Africa, which is home to almost 24.5 million people with the virus, or 64% of all people living with HIV. UNAIDS estimates that 2.7 million became newly infected in this region in 2005 (31). A considerable number of new HIV infections are transmitted by recent HIV seroconverters (6, 23, 25, 26).Early diagnosis of HIV infection is important, as it allows appropriate clinical management and counseling of patients in order to prevent further sexual transmission of HIV to partners (11, 24). It is also important in special situations, for example, screening blood for transfusion (10, 19). Access to reliable test systems for diagnosis of HIV infection at the earliest possible stage is necessary for effective prevention of transmission, early intervention, access to antiviral therapy, surveillance, and blood safety.The enzyme-linked immunospot assay is a well-established method for the enumeration of antibody-secreting cells and study of spontaneous secretion of antibodies (4, 7, 8, 12). In this study, we utilized the PlasmAcute technology to detect anti-HIV-1 antibodies in B-cell lysates before they are detected in serum. This technology is based on the observation that B cells from peripheral blood contain functional antibodies elicited by an infectious agent or vaccine, and these antibodies can be measured before they can be detected in plasma (14, 22). This allows a narrowing of the “window period” between immunological stimulation and seroconversion. The technology involves initial capture enrichment and isolation of B cells from a peripheral-whole-blood sample by using paramagnetic polystyrene beads coated with monoclonal anti-CD19 antibodies followed by subsequent lysis and testing of the lysate in an appropriate immunoassay (22).(The data in this paper were presented in put at the joint meeting of the 17th Meeting of the International Society for Sexually Transmitted Diseases Research and the 10th World Congress of the International Society against Sexually Transmitted Infections, Seattle, WA, 29 July to 1 August 2007 [26a].)     

Quantitation of Human Immunodeficiency Virus Type 2 DNA in Peripheral Blood Mononuclear Cells by Using a Quantitative-Competitive PCR Assay

A new quantitative-competitive PCR-based human immunodeficiency virus type 2 (HIV-2) proviral DNA assay (QC-PCR) was developed and used to determine the proviral load in HIV-2-infected individuals. Proviral load varied considerably, with means of 1,831 copies per 10⁶ peripheral blood mononuclear cells for asymptomatic subjects (n = 19) and 2,587 for AIDS patients (n = 2). HIV-2 viral and proviral loads also varied significantly over time in asymptomatic patients. These data suggest that a high level of virus replication occurs throughout the asymptomatic phase of HIV-2 infection.     

Detection of Antibodies to Varicella-Zoster Virus in Recipients of the Varicella Vaccine by Using a Luciferase Immunoprecipitation System Assay

A high-throughput test to detect varicella-zoster virus (VZV) antibodies in varicella vaccine recipients is not currently available. One of the most sensitive tests for detecting VZV antibodies after vaccination is the fluorescent antibody to membrane antigen (FAMA) test. Unfortunately, this test is labor-intensive, somewhat subjective to read, and not commercially available. Therefore, we developed a highly quantitative and high-throughput luciferase immunoprecipitation system (LIPS) assay to detect antibody to VZV glycoprotein E (gE). Tests of children who received the varicella vaccine showed that the gE LIPS assay had 90% sensitivity and 70% specificity, a viral capsid antigen enzyme-linked immunosorbent assay (ELISA) had 67% and 87% specificity, and a glycoprotein ELISA (not commercially available in the United States) had 94% sensitivity and 74% specificity compared with the FAMA test. The rates of antibody detection by the gE LIPS and glycoprotein ELISA were not statistically different. Therefore, the gE LIPS assay may be useful for detecting VZV antibodies in varicella vaccine recipients. (This study has been registered at ClinicalTrials.gov under registration no. {"type":"clinical-trial","attrs":{"text":"NCT00921999","term_id":"NCT00921999"}}NCT00921999.)