Clinical Evaluation of BioPlex 2200 HIV Ag-Ab,an Automated Screening Method Providing Discrete Detection of HIV-1 p24 Antigen,HIV-1 Antibody,and HIV-2 Antibody

Early and accurate diagnosis is essential for optimal therapeutic outcomes in patients infected with HIV. Currently, none of the commercially available fourth-generation assays differentiate HIV-1 and HIV-2 antibodies (Ab) or the HIV-1 p24 antigen (Ag). The aim of this study was to evaluate the performance of a novel assay, the BioPlex 2200 HIV Ag-Ab. This assay uses a multiplex flow immunoassay design allowing the simultaneous detection and identification of antibodies to HIV-1 (groups M and O), HIV-2, and the HIV-1 p24 antigen, in addition to providing a traditional composite result. A total of 1,505 routine serum samples were prospectively tested. Results were compared with those from the Architect HIV Combo assay. The sensitivity of the BioPlex 2200 was 100%. The specificity assessed on repeated false-positive samples was 99.5%. In addition, 524 frozen specimens from patients known to be infected with HIV-1 or HIV-2 were tested. Of these specimens, 420 were infected with HIV-1, including 156 of known genotypes, 86 were infected with HIV-2, 7 were infected with HIV-1 and HIV-2, and 11 were from patients with acute HIV infection. Sensitivity was 100% for the HIV genotypes tested. The differentiation capabilities of the BioPlex 2200 HIV Ag-Ab assay for HIV-1, HIV-2, dual HIV-1/HIV-2, and early infections were 100%, 90.7%, 100%, and 90.9%, respectively. The BioPlex 2200 is a sensitive and specific assay that offers advantages over conventional HIV combo assays, also referred to as fourth-generation assays, to accurately differentiate and report HIV-1 p24 antigen and HIV-1 and HIV-2 antibodies.     

Comparison of the clinical performance of the ECLusys HIV combi assay with the Lumipulse f and HISCL 2000-i HIV-1/2 ab screening assays

We compared the ECLusys HIV combi assay (ECL HIV Ag/Ab) to the Lumipulse Forte (LPf HIV 1/2 Ab) and HISCL (HIS HIV 1/2 Ab) assays. In a dilution sensitivity test using dilution panels of WHO HIV antibody international reference panel (HIV-1 Subtype A, B, C, E, HIV-1 Group O, HIV-2) and HIV-1/2 Ab CE marked material(HIV-1, HIV-2) parent specimens, the ECL assay enabled detection at a higher level of sensitivity than either the LPf assay or the HIS assay for all dilution panels. In an early detection test in the early phase of infection in which a BBI HIV seroconversion panel was used, the ECL assay enabled detection 7 days after initial blood sample collection, whereas the LPf and HIS assays enabled detection after 27 days. In a specificity test using high RF positive specimens (n=33), pregnancy specimens (n=35), cytomegalovirus antibody positive specimens (n=36), and high M protein positive specimens (n=21) that were confirmed negative for HIV-1/2 antibodies by the LPf assay, negative results were obtained for all specimens on both the ECL assay and the HIS assay. In a correlation test using routinely collected clinical specimens (n=121), including positive stock specimens, the ECL and HIS assays demonstrated the highest agreement rate 98.3%. The above results confirmed that the fourth-generation reagent ECL assay, which simultaneously detects both HIV-1/2 antibodies and p24 antigens, is both highly sensitive and specific, and is a suitable assay for use in routine testing.     

Diagnostic challenges for rapid human immunodeficiency virus assays. Performance using HIV-1 group O, HIV-1 group M, and HIV-2 samples

OBJECTIVE: We sought to determine the ability of seven rapid assays for human immunodeficiency virus (HIV) to detect antibodies in a panel of sera from individuals infected with different types and groups of HIV. STUDY DESIGN/METHODS: Sixty-eight well-characterized samples, including HIV-1 group O (24), several HIV-1 group M clades (21), HIV-1/2 (10), HIV-2 (10), and samples with indeterminate results (3), were tested by the following rapid HIV assays: HIV-Spot, HIVCHEK System 3, A/Q Rapid HIV, Genie II HIV-1/HIV-2, Quix HIV-1-2-O, ImmunoComb II HIV-1+2 BiSpot, and the Serodia HIV-1+2. RESULTS: All tests successfully detected the HIV-1 group M clades and the HIV-1/2-positive samples. Of the HIV-2 stand-alone samples, four tests missed the same sample, and three tests missed another sample. Of the HIV-1 group O samples, four samples were missed by at least one test, and another sample was missed by three tests. The sensitivity of the seven rapid assays in detecting each group of sera was between 83% and 100%, with only one test having a sensitivity of 100% for all groups of sera. Three samples proved to be problematic because they were misclassified by more than one assay. CONCLUSIONS: The performance of rapid HIV assays is variable when testing sera from individuals infected with HIV-1 group O and HIV-2.     

Rapid Assay for Simultaneous Detection and Differentiation of Immunoglobulin G Antibodies to Human Immunodeficiency Virus Type 1 (HIV-1) Group M, HIV-1 Group O, and HIV-2

A rapid immunodiagnostic test that detects and discriminates human immunodeficiency virus (HIV) infections on the basis of viral type, HIV type 1 (HIV-1) group M, HIV-1 group O, or HIV-2, was developed. The rapid assay for the detection of HIV (HIV rapid assay) was designed as an instrument-free chromatographic immunoassay that detects immunoglobulin G (IgG) antibodies to HIV. To assess the performance of the HIV rapid assay, 470 HIV-positive plasma samples were tested by PCR and/or Western blotting to confirm the genotype of the infecting virus. These samples were infected with strains that represented a wide variety of HIV strains including HIV-1 group M (subtypes A through G), HIV-1 group O, and HIV-2 (subtypes A and B). The results showed that the HIV genotype identity established by the rapid assay reliably (469 of 470 samples) correlates with the HIV genotype identity established by PCR or Western blotting. A total of 879 plasma samples were tested for IgG to HIV by a licensed enzyme immunoassay (EIA) (470 HIV-positive samples and 409 HIV-negative samples). When they were tested by the rapid assay, 469 samples were positive and 410 were negative (99.88% agreement). Twelve seroconversion panels were tested by both the rapid assay and a licensed EIA. For nine panels identical results were obtained by the two assays. For the remaining three panels, the rapid assay was positive one bleed later in comparison to the bleed at which the EIA was positive. One hundred three urine samples, including 93 urine samples from HIV-seropositive individuals and 10 urine samples from seronegative individuals, were tested by the rapid assay. Ninety-one of the ninety-three urine samples from HIV-seropositive individuals were found to be positive by the rapid assay. There were no false-positive results (98.05% agreement). Virus in all urine samples tested were typed as HIV-1 group M. These results suggest that a rapid assay based on the detection of IgG specific for selected transmembrane HIV antigens provides a simple and reliable test that is capable of distinguishing HIV infections on the basis of viral type.     

Fine serotyping of human immunodeficiency virus serotype 1 (HIV-1) and HIV-2 infections by using synthetic oligopeptides representing an immunodominant domain of HIV-1 and HIV-2/simian immunodeficiency virus.

In this study, enzyme immunoassays for detection of type-specific antibodies to human immunodeficiency viruses (HIV) were developed by using short peptides corresponding to sequences located within the immunodominant domain of the transmembrane glycoproteins of both HIV-1 and HIV-2-simian immunodeficiency virus (SIV). The assays were highly sensitive with currently available sera from various geographical areas. Furthermore, they appeared to be more specific in HIV serotyping than the Western blot (immunoblot) assay, since all of the sera were clearly discriminated as one or the other type. It was also shown that in contrast to HIV-1, the C-terminal cysteine residue (amino acid 620, SIV from captive macaques, Mm142 strain) of the HIV-2-SIV peptide is not necessary for recognition of the peptide by antibody to HIV-2.     

Evaluation of three rapid tests for detection of antibodies to HIV-1 non-B subtypes

Three rapid tests were evaluated for the detection of antibodies to HIV in 111 plasma specimens (81 HIV-1 non-B subtypes, including 4 coinfections with HIV-2, 30 HIV negative). The Determine HIV-1/2, Equipar HIV, and Multispot HIV-1/2 tests were 100% sensitive and specific for the detection of HIV-1 antibodies. Multispot showed 92.7% specificity for HIV-2.     

Serological detection of infection with diverse human and simian immunodeficiency viruses using consensus env peptides

Cross-species transmission has been shown to play an important role in the emergence of human retroviruses. We developed a generic enzyme immunoassay using synthetic peptides from gp41 and C2V3 consensus sequences (human immunodeficiency virus [HIV] type 1 [HIV-1] groups M, O, and N and the homologous region of simian immunodeficiency virus [SIV] strains from chimpanzees [SIVcpz], SIVcpzGAB1 and SIVcpzANT) to detect divergent HIV and SIV. A cocktail of peptides from gp41 and C2V3 (M-O) detected all HIV-1 group M and O sera and showed cross-reactivity with SIVcpz sera. Further, a mixture of C2V3 peptides (GAB1-ANT) failed to detect HIV-1 infections but reacted with all SIVcpz sera, allowing discrimination of SIVcpz from HIV-1 infections. Since most SIVcpz sera cross-reacted with HIV-1 peptides, we next evaluated SIVcpz serum reactivity with rapid tests for HIV-1/2. SIVcpzANT and SIVcpzUS sera reacted with the Sero-strip and Multispot assays. Both tests are sensitive in detecting group M (97 100%, respectively), although Multispot has lower sensitivity for group O detection (67%) than does Sero-strip (100%). The limited volume and time required to perform these assays make them a generic tool for field screening. The env peptide-based assay and rapid tests should allow for the identification of emerging variants of HIV and SIV.     

Comparing modified and plain peptide linked enzyme immunosorbent assay (ELISA) for detection of human immunodeficiency virus type-1 (HIV-1) and type-2 (HIV-2) antibodies

Serological diagnosis of human immunodeficiency virus (HIV) based on detection of HIV antibodies is one of the easiest, cheapest and simplest assay. Synthetic peptides corresponding to immunodominant regions of envelope glycoprotein (gp41, V3 loop for HIV-1 and gp36 for HIV-2) were used in the present study, to detect the anti-HIV antibodies in sera of Sexually Transmitted Diseases (STD), Tuberculosis (TB), Anti-Natal Care (ANC) patients. About 550 serum samples were tested using Enzyme Linked Immunosorbent Assay (ELISA) technique. The human sera positive for antibody to HIV-1 and HIV-2, reacted to different degrees with these peptides when used as a plain peptide with or without CGG motif/biotin motif at the amino terminus. The selected sequences are of Indian strain with 'C' serotype. The results showed a 100% sensitivity and specificity for V3 loop peptide and 98% sensitivity and specificity for gp41 peptide containing CGG moiety while the plain peptides showed similar sensitivities but low specificity's, i.e. 98% for V3 loop peptide and 42% for gp41 peptide when reacted with HIV-1 positive sera. The presence of biotin at the amino terminus did not provide any beneficial effect in increasing the sensitivity although the specificity was enhanced for both the peptide sequences, i.e. gp41 and V3 loop peptide. Furthermore, the gp36 peptide containing CGG moiety detected the HIV-2 sera with 100% sensitivity and 98% specificity while the sensitivity and specificity of gp36 plain peptide was reduced to 98 and 90%. Thus the study overall highlighted the importance of synthetic peptides containing CGG moiety as a capture antigen in detecting both HIV-1 & 2 sera using an indigenously built ELISA system which is simple, cheap, sensitive and cost effective for rural areas.     

Development and evaluation of HIV-1 subtype RNA panels for the standardization of HIV-1 NAT assays

Multiple nucleic acid-based techniques (NAT) have been implemented for testing blood and plasma donors for HIV-1 RNA which may be detected at an earlier stage of infection when HIV antigen or antibody is absent or below the limit of detection of current assays. The available NAT assays are based on different technologies. In order to evaluate the performance of nucleic acid-based techniques (NAT assays) and to allow accurate comparisons of results from different assays, it is essential to have well characterized specimens with known copy numbers as a standard. For this purpose, a comprehensive study was conducted to develop two HIV-1 RNA reference panels. The first (Panel 1) was prepared using a single specimen from the HIV-1 group M subtype B and consists of panel members with a wide range of HIV-1 RNA copy numbers. Panel 2 consists of 26 members representing HIV-1 group M subtypes A, C, D, E, F, G and groups O and N. For accurate determination of HIV-1 RNA copy numbers of each member of Panel 2, they were analyzed using various testing platforms/technologies available through the cooperation of five independent laboratories participating in the study. A consensus value for HIV RNA copy number was assigned to each member of Panel 2 based on statistical analysis of the data provided by the participants. Both panels could serve as reference panels to be used by manufacturers of HIV NAT tests to evaluate the sensitivity limits of their assays.     

Increased immunoglobulin G, but not M, binding to endogenous retroviral antigens in HIV-1 infected persons

The modes of interaction between products of human endogenous retroviral (HERV) sequences and the immune system are largely unknown. In HIV infected persons, an exogenous retrovirus adds further complexity to the situation. Therefore, 14 synthetic peptides with sequences derived from conserved regions of various endogenous retroviruses (ERVs) and from related exogenous retroviruses were used to search for IgG and IgM antibodies that bind to such antigens in 15 HIV-1 seropositive and 17 seronegative immunosuppressed patients. IgG binding to three peptides, namely, the C-terminal half of murine leukemia virus (MLV) capsid protein, the conserved portion of HERV-H transmembrane protein, and the Pol region of human mouse mammary tumor virus (MMTV)-like (HML3) sequence, was observed in both groups. Binding was, however, more frequent and more firm in HIV-1 positive samples (P<0.0001, Wilcoxon rank sum test). IgM binding to the same peptides showed no significant differentiation between the two groups of patients. Binding to both immunoglobulin isotypes was sometimes variable over time in both groups. No correlation of either IgG or IgM peptide binding with progression to AIDS in HIV-1 infected individuals was observed. Inhibition studies using analogous endogenous and exogenous retroviral peptides, including HIV-1, demonstrated specificity of the IgG antibodies for a narrow range of MLV- and MMTV-like retroviral antigens, and excluded cross-reactivity of antibodies to HIV-1 as a cause of these observations. Thus, unlike IgG, IgM binding to retroviral antigens was ubiquitous. It is suggested that anti-HERV IgM belong to a class of natural antibodies and might serve as primers in the mediation of humoral immune responses to more or less related exogenous retroviruses. Increased IgG binding in HIV-1 infected individuals could result from such priming, or reflect higher HERV antigen expression.     

Seven human immunodeficiency virus (HIV) antigen-antibody combination assays: evaluation of HIV seroconversion sensitivity and subtype detection

In this study, we evaluated the performance of two prototype human immunodeficiency virus (HIV) antigen-antibody (Ag-Ab) combination assays, one from Abbott Laboratories (AxSYM HIV Ag-Ab) and the other from bioMerieux (VIDAS HIV Duo Ultra), versus five combination assays commercially available in Europe. The assays were Enzygnost HIV Integral, Genscreen Plus HIV Ag-Ab, Murex HIV Ag-Ab Combination, VIDAS HIV Duo, and Vironostika HIV Uniform II Ag-Ab. All assays were evaluated for the ability to detect p24 antigen from HIV-1 groups M and O, antibody-positive plasma samples from HIV-1 groups M and O, HIV-2, and 19 HIV seroconversion panels. Results indicate that although all combination assays can detect antibodies to HIV-1, group M, subtypes A to G, circulating recombinant form (CRF) A/E, and HIV-1 group O, their sensitivity varied considerably when tested using diluted HIV-1 group O and HIV-2 antibody-positive samples. Among combination assays, the AxSYM, Murex, and VIDAS HIV Duo Ultra assays exhibited the best antigen sensitivity (at approximately 25 pg of HIV Ag/ml) for detection of HIV-1 group M, subtypes A to G and CRF A/E, and HIV-1 group O isolates. However, the VIDAS HIV Duo Ultra assay had a lower sensitivity for HIV-1 group M and subtype C, and was unable to detect subtype C antigen even at 125 pg of HIV Ag/ml. The HIV antigen sensitivity of the VIDAS HIV Duo and Genscreen Plus combination assays was approximately 125 pg of HIV Ag/ml for detection of all HIV-1 group M isolates except HIV-1 group O while the sensitivity of Vironostika HIV Uniform II Ag-Ab and Enzygnost HIV Integral Ag-Ab assays for all the group M subtypes was >125 pg of HIV Ag/ml. Among the combination assays, the AxSYM assay had the best performance for detection of early seroconversion samples, followed by the Murex and VIDAS HIV Duo Ultra assays.     

Analysis of genetic variability within the immunodominant epitopes of envelope gp41 from human immunodeficiency virus type 1 (HIV-1) group M and its impact on HIV-1 antibody detection

The serodiagnosis of human immunodeficiency virus type 1 (HIV-1) infection primarily relies on the detection of antibodies, most of which are directed against the immunodominant regions (IDR) of HIV-1 structural proteins. Among these, the N-terminal region of gp41 contains cluster I (amino acids [aa] 580 to 623), comprising the cytotoxic T-lymphocyte epitope (AVERYLKDQQLL) and the cysteine loop (CSGKLIC), and cluster II (aa 646 to 682), comprising an ectodomain region (ELDKWA). To delineate the epitope diversity within clusters I and II and to determine whether the diversity affects serologic detection by U.S. Food and Drug Administration (FDA)-licensed enzyme immunoassay (EIA) kits, gp41 Env sequences from 247 seropositive persons infected with HIV-1 group M, subtypes A (n = 42), B (n = 62), B' (n = 13), C (n = 38), D (n = 41), E (n = 18), F (n = 27), and G (n = 6), and 6 HIV-1-infected but persistently seronegative (HIPS) persons were analyzed. While all IDR were highly conserved among both seropositive and HIPS persons, minor amino acid substitutions (<20% for any one residue, mostly conservative) were observed for all subtypes, except for B', in comparison with the consensus sequence for each subtype. Most importantly, none of the observed substitutions among the group M plasma specimens affected antibody detection, since all specimens (n = 152) tested positive with all five FDA-licensed EIA kits. Furthermore, all specimens reacted with a group M consensus gp41 peptide (WGIKQLQARVLAVERYLKDQQLLGIWGCSGKLICTTAVPWNASW), and high degrees of cross-reactivity (>80%) were observed with an HIV-1 group N peptide, an HIV-1 group O peptide, and a peptide derived from the homologous region of gp41 from simian immunodeficiency virus from chimpanzee (SIVcpz). Taken together, these data indicate that the minor substitutions observed within the IDR of gp41 of HIV-1 group M subtypes do not affect antibody recognition and that all HIV-1-seropositive specimens containing the observed substitutions react with the FDA-licensed EIA kits regardless of viral genotype and geographic origin.     

Serological Detection of Infection with Diverse Human and Simian Immunodeficiency Viruses Using Consensus env Peptides

Cross-species transmission has been shown to play an important role in the emergence of human retroviruses. We developed a generic enzyme immunoassay using synthetic peptides from gp41 and C2V3 consensus sequences (human immunodeficiency virus [HIV] type 1 [HIV-1] groups M, O, and N and the homologous region of simian immunodeficiency virus [SIV] strains from chimpanzees [SIVcpz], SIVcpzGAB1 and SIVcpzANT) to detect divergent HIV and SIV. A cocktail of peptides from gp41 and C2V3 (M-O) detected all HIV-1 group M and O sera and showed cross-reactivity with SIVcpz sera. Further, a mixture of C2V3 peptides (GAB1-ANT) failed to detect HIV-1 infections but reacted with all SIVcpz sera, allowing discrimination of SIVcpz from HIV-1 infections. Since most SIVcpz sera cross-reacted with HIV-1 peptides, we next evaluated SIVcpz serum reactivity with rapid tests for HIV-1/2. SIVcpzANT and SIVcpzUS sera reacted with the Sero-strip and Multispot assays. Both tests are sensitive in detecting group M (97 100%, respectively), although Multispot has lower sensitivity for group O detection (67%) than does Sero-strip (100%). The limited volume and time required to perform these assays make them a generic tool for field screening. The env peptide-based assay and rapid tests should allow for the identification of emerging variants of HIV and SIV.     

Monoclonal antibodies to an HIV-1 group O envelope recombinant

Monoclonal antibodies were developed to a recombinant HIV-I group O envelope protein derived from the isolate HAM112. These monoclonal antibodies were characterized for reactivity to a series of overlapping synthetic peptides (29-30 mers) covering gp120 C-terminal and gp41 ectodomain regions of the HIV-1 group O envelope protein. Most of these monoclonal antibodies reacted with peptides spanning sequences analogous to HIV-1 group M epitopes identified from studies in mice and humans. However, several of the antibodies that were nonreactive to individual peptides did react to a mixture of longer peptides from the N-terminal and C-terminal helical regions of the gp41 ectodomain. The monoclonal antibodies described in this study are valuable tools for characterization of antigenic differences between HIV-1 group O and group M viruses.     

Multicenter evaluation of a new, automated enzyme-linked immunoassay for detection of human immunodeficiency virus-specific antibodies and antigen

A collaborative multicenter study was conducted to evaluate the sensitivity, specificity, and precision of a three-step, fully automated, qualitative microparticle-based enzyme-linked immunoassay (AxSYM HIV Ag/Ab Combo; Abbott Laboratories), designed to simultaneously detect (i). antibodies against human immunodeficiency virus type 1 (HIV-1) and/or type 2 (HIV-2) and (ii). HIV p24 antigen. A significant reduction in the HIV seroconversion window was achieved by combining detection of HIV antibodies and antigen into a single assay format. For 22 selected, commercial HIV seroconversion panels, the mean time of detection with the combined-format HIV antigen-antibody assay was reduced by 6.15 days compared to that with a similar third-generation single-format HIV antibody assay. The quantitative sensitivity of the combination assay for the p24 antigen (17.5 pg/ml by use of the p24 quantitative panel VIH SFTS96') was nearly equivalent to that of single-format antigen tests. The combination assay demonstrated sensitive (100%) detection of anti-HIV immunoglobulin in specimens from individuals in CDC stages A, B, and C and from individuals infected with different HIV-1 group M subtypes, group O, or HIV-2. The apparent specificity for hospitalized patients (n = 1938) was 99.90%. In a random population of 7900 volunteer blood donors, the specificity (99.87%) was comparable to that of a third-generation single-format HIV antibody assay (99.92%) on the same donor specimens. In addition, the combination assay was robust to potential interfering specimens. The precision of the combination was high, with intra- and interrun variances of 相似文献   

Impact of human immunodeficiency virus type 1 (HIV-1) genetic diversity on performance of four commercial viral load assays: LCx HIV RNA Quantitative, AMPLICOR HIV-1 MONITOR v1.5, VERSANT HIV-1 RNA 3.0, and NucliSens HIV-1 QT

Human immunodeficiency virus type 1 (HIV-1) evolution and changing strain distribution present a challenge to nucleic acid-based assays. Reliable patient monitoring of viral loads requires the detection and accurate quantification of genetically diverse HIV-1. A panel of 97 HIV-1-seropositive plasma samples collected from Cameroon, Brazil, and South Africa was used to compare the performance of four commercially available HIV RNA quantitative tests: Abbott LCx HIV RNA Quantitative assay (LCx), Bayer Versant HIV-1 RNA 3.0 (bDNA), Roche AMPLICOR HIV-1 MONITOR v1.5 (Monitor v1.5), and bioMérieux NucliSens HIV-1 QT (NucliSens). The panel included group M, group O, and recombinant viruses based on sequence analysis of gag p24, pol integrase, and env gp41. The LCx HIV assay quantified viral RNA in 97 (100%) of the samples. In comparison, bDNA, Monitor v1.5, and NucliSens quantified viral RNA in 96.9%, 94.8%, and 88.6% of the samples, respectively. The two group O specimens were quantified only by the LCx HIV assay. Analysis of nucleotide mismatches at the primer/probe binding sites for Monitor v1.5, NucliSens, and LCx assays revealed that performance characteristics reflected differences in the level of genetic conservation within the target regions.     

Salivary detection of HIV-1 antibodies using recombinant HIV-1 peptides

Salivary antibodies may play a role in the absence of HIV-1 transmission by saliva. We evaluated the presence of salivary IgG antibodies to HIV-1 using a recombinant ELISA. Whole saliva was collected from 21 HIV-1-seropositive individuals and assayed in an ELISA, ASQ (Beckman Instruments, Brea, CA), consisting of a panel of six HIV-1 recombinant peptides. Saliva samples from 20 individuals demonstrated IgG to one or more peptides and 18 to two or more peptides. Samples from 20 seropositive individuals were reactive with the gp41 peptide, whereas only 12 were reactive with the two gp120 peptides. Nineteen of twenty salivas also had detectable IgG antibodies to HIV-1 by Western blotting. The results indicate that viral-specific IgG antibodies are present in the saliva of a high percentage of HIV-infected individuals and that a recombinant peptide ELISA for saliva might be useful for the detection of HIV-1 infection.     

A new antigen scanning strategy for monitoring HIV-1 specific T-cell immune responses

Delineation of the immune correlates of protection in natural infection or after vaccination is a mandatory step for vaccine development. Although the most recent techniques allow a sensitive and specific detection of the cellular immune response, a consensus on the best strategy to assess their magnitude and breadth is yet to be reached. Within the AIDS Vaccine Integrated Project (AVIP http://www.avip-eu.org) we developed an antigen scanning strategy combining the empirical-based approach of overlapping peptides with a vast array of database information. This new system, termed Variable Overlapping Peptide Scanning Design (VOPSD), was used for preparing two peptide sets encompassing the candidate HIV-1 vaccine antigens Tat and Nef. Validation of the VOPSD strategy was obtained by direct comparison with 15mer or 20mer peptide sets in a trial involving six laboratories of the AVIP consortium. Cross-reactive background responses were measured in 80 HIV seronegative donors (HIV-), while sensitivity and magnitude of Tat and Nef-specific T-cell responses were assessed on 90 HIV+ individuals. In HIV-, VOPSD peptides generated background responses comparable with those of the standard sets. In HIV-1+ individuals the VOPSD pools showed a higher sensitivity in detecting individual responses (Tat VOPSD vs. Tat 15mers or 20mers: p≤0.01) as well as in generating stronger responses (Nef VOPSD vs. Nef 20mers: p<0.001) than standard sets, enhancing both CD4 and CD8 T-cell responses. Moreover, this peptide design allowed a marked reduction of the peptides number, representing a powerful tool for investigating novel HIV-1 candidate vaccine antigens in cohorts of HIV-seronegative and seropositive individuals.     

Quantification of HIV-1 group M (subtypes A-G) and group O by the LCx HIV RNA quantitative assay

Human immunodeficiency virus type 1 (HIV-1) genetic diversity presents a challenge to nucleic acid-based assays with regard to sensitivity of detection and accuracy of quantification. The Abbott LCx HIV RNA Quantitative assay (LCx(R) HIV assay), a competitive RT-PCR targeting the pol integrase region, was evaluated using a panel of 297 HIV-1 seropositive plasma samples from Cameroon, Uganda, Brazil, Thailand, Spain, Argentina and South Africa. The panel included group M subtypes A-G, mosaics, and group O based on sequence analysis of gag p24, pol integrase, and env gp41. The LCx HIV assay quantified 290 (97.6%) of the samples, including all the group O samples tested. In comparison, the Roche AMPLICOR HIV-1 MONITOR test versions 1.0 and 1.5 quantified 67.3 and 94.6% of the samples, respectively. No group O specimens were quantified by either version of AMPLICOR HIV-1 MONITOR. Seven specimens were below the detectable limits of all the three assays. The LCx HIV assay had fewer nucleotide mismatches at primer/probe binding sites as compared with both AMPLICOR HIV-1 MONITOR tests. The high degree of nucleotide conservation within the pol target region enables the LCx HIV assay to efficiently quantify the HIV-1 subtypes A-G and the most genetically diverse HIV-1, group O.