OBJECTIVETo describe and evaluate a double-antigen sandwich ELISA for detecting human immunodeficiency virus type 1/2 (HIV-1/2) specific antibodies.

METHODSThe peptides gp41.1(sp1), gp41.2(sp2), gp120(sp3) and p24(sp4) of HIV-1 and gp36(sp5) of HIV-2 were artificially synthesized. Then sp1, sp3, sp4 and sp5 were used as coating antigens; sp1, sp2, sp4 and sp5 labeled with HRP were used as conjugates in this sandwich ELISA.

RESULTSThe specificity and sensitivity of the assay were both 100% in detecting anti-HIV of 40 control sera of the second generation panel, higher than indirect ELISA (specificity 90% and sensitivity, 65%, respectively). All of 210 sera from individuals with other diseases were negative for anti-HIV. The consistency rate was 100% when our sandwich ELISA and Abbott HIVAB were used to detect anti-HIV in 90 healthy blood donors and 88 HIV infected individuals.

CONCLUSIONSThe results showed that this sandwich ELISA for detection of anti-HIV is specific, sensitive and convenient, and it is suitable for screening blood donors and detecting HIV infection.

Enzyme-Linked Immunosorbent Assay ; methods ; HIV Antibodies ; blood ; HIV Infections ; blood ; virology ; HIV-1 ; immunology ; HIV-2 ; immunology ; Humans

OBJECTIVETo explore the follow-up visit, outcome and auxiliary diagnosis method on the cases with indeterminate antibody level measured by Western blotting as well as the related biological factors.

METHODSThe cases with indeterminate result were followed up according to the National Guideline for Detection of HIV/AIDS (2009) and samples were collected for HIV antibody detection, p24 antigen and nucleic acid were detected as a supplementary diagnosis at the same time. The samples were also be detected for HBV, HCV, TP, HTLV-I/II, ANA, and AFP, and the results were compared to that of screened positive and confirmed negative cases.

RESULTSA total of 73 were followed up successfully and taken a second HIV test, 25 cases were tested positive and 48 were tested negative for HIV during the follow-up period. For the 25 HIV positive cases, the HIV seroconversion rate was 100.00% at any time point when the interval between the first and returning detection was longer than 1 week. The major Western blotting bands for the cases with indeterminate result were p24 and gp160 and it was different between HIV positive and negative cases in Western blotting band profiles. The consistency and sensitivity of nucleic acid detection were higher than 90.00%, and were higher than that of p24 antigen (69.09% (38/55) and 27.27% (6/22)) (χ(2)(consistency) = 6.875, χ(2)(sensitivity) = 18.893, P < 0.05). The positive rates of ANA and AFP of indeterminate cases excluded from HIV infection were 20.83% (10/28) and 6.25% (3/48) and higher than that of screened positive and confirmed negative cases (0.00%), the difference had statistic significance (χ(2)(ANA) = 19.430, χ(2)(AFP) = 5.520, P < 0.05).

CONCLUSIONIt is critical to get timely diagnosis for the indeterminate cases according to the new national guideline for detection of HIV/AIDS. Nucleic acid detection has higher application value as auxiliary diagnosis for HIV infection than p24 antigen. The increased levels of ANA and AFP may be the factors resulting in the nonspecific indeterminate results.

Antibodies, Antinuclear ; blood ; Female ; Follow-Up Studies ; HIV Antibodies ; blood ; HIV Infections ; diagnosis ; immunology ; Humans ; Male ; alpha-Fetoproteins ; analysis

BACKGROUND: Many immunochromatography (ICA) kits for anti-human immunodeficiency virus type (HIV) antibody (Ab) have been introduced to improve the accessibility of HIV Ab tests. However, qualified evaluation reports for HIV rapid tests are not enough to validate their performances. Metaanalysis for the performances of the HIV Ab rapid tests was performed in this study. METHODS: PubMed database was searched with combination of search terms, 'human immunodeficiency virus', 'HIV Ab', 'rapid test', 'immunochromatography', 'performance', 'sensitivity', and 'specificity'. Criteria of inclusion were performance studies for HIV ICA kits with serum or EDTA whole blood. Methodological qualities were evaluated with standards for reporting of diagnostic accuracy studies (STARD) checklists by two investigators. Homogeneity among selected studies was evaluated and then pooled sensitivity and specificity were calculated. Positive and negative predictive values were simulated with presumed HIV prevalence in Korea. RESULTS: Twenty-three studies were selected from 12 high-qualified papers with STARD checklists. The performance of 23 studies were found to be heterogeneous (P<0.1) and random effect model was used. Pooled sensitivity was 99.71% (95% CI: 99.45-99.97%) and pooled specificity was 99.27% (95% CI: 98.83-99.70%). With HIV prevalence of 0.03%, positive and negative predictive values were presumed to be 3.936% and 99.999%, respectively. CONCLUSIONS: This meta-analysis for HIV ICA rapid tests showed good performance. In consideration of low positive predictive values of HIV rapid tests, confirmation by enzyme immunoassay or Western blot is still needed. This study would be helpful in evaluating and establishing proper performance guideline for those kits not fully evaluated.
HIV Antibodies/*blood/immunology ; HIV Infections/*diagnosis ; Humans ; Reagent Kits, Diagnostic ; Sensitivity and Specificity

OBJECTIVETo establish and evaluate an Enzyme Immunoassay diagnostic kit combined with anti-HIV1/2 antibody and P24 antigen for shortening the examination window period of HIV infection in HIV laboratory diagnosis.

METHODSThe enzyme-linked reaction plates was coated by anti-HIV P24 monoclonal antibody and HIV 1/2 antigen. Labeling HIV1/2 antigen and anti-HIV P24 polyclonal antibody with horseradish peroxidase, setup an integrated ELISA kit for detecting anti-HIV-1/2 antibody and HIV P24 antigen, and evaluate the specificity and sensitivity of this kit.

RESULTSThe sensitivity of testing P24 antigen was up to 0.2 ng/ml. 78 serum samples of patients with AIDS, 85 serum samples of healthy people were compared with Abbott EIA kit, the coincidence was 100%. 12 051 sera from normal persons and patients were examined, the sensitivity of 100 %and specificity of 99.62 %, respectively.

CONCLUSIONThe anti-HIV1/2 antibody and HIV P24 antigen can be measured at the same time using this EIA kit, while the examination window period of HIV infection is shortened. Thus, the method is suitable for laboratory diagnosis and epidemiological investigation.

Enzyme-Linked Immunosorbent Assay ; HIV Antibodies ; blood ; HIV Core Protein p24 ; blood ; HIV-1 ; immunology ; HIV-2 ; immunology ; Humans ; Reagent Kits, Diagnostic

OBJECTIVETo study the incidence and risk factors of HIV and syphilis seroconversion among men who have sex with men (MSM) in Beijing.

METHODSA total of 550 MSM were recruited on the basis of community and followed up after 6 and 12 months in Beijing. Each subject was investigated by only one investigator at one time to collect information on demographics and behaviors. Blood samples were collected to test HIV and syphilis seroconversion. ELISA was used for screening test, west blotting (WB) and Particle agglutination were used for confirmatory test.

RESULTSA total of 550 MSM investigated, among which 4.5% (25/550) were HIV-positive and 29.3% (161/550) were syphilis-positive. For 525 HIV-negative MSM, 87.0% (457/525) retained during the 12-month investigation. Seroincidence for HIV and syphilis were 3.37/100 person-years (95%CI = 1.66 - 5.08) and 9.32/100 person-years (95%CI = 5.87 - 12.77) respectively. HIV seroconversions for those who performed and did not perform rectal douching after homosexual anal intercourse in the past 3 months were 7.11/100 and 0.76/100 person-years respectively. Multivariate Cox regression analysis revealed that rectal douching after homosexual anal intercourse in the past 3 months (HR = 9.23, 95%CI = 2.08 - 40.88) was significantly associated with HIV seroconversion. Syphilis seroconversions for those who met male sex partners in parks, public washrooms or bathhouses in the past 3 months were 41.77/100 and 7.97/100 person-years respectively. Syphilis seroconversions for those who performed and did not perform rectal douching after homosexual anal intercourse in the past 3 months were 16.17/100 and 4.92/100 person-years respectively. In the past 3 months, meeting male sex partners in parks, public washrooms or bathhouses (HR = 4.67, 95%CI = 1.77 - 12.34) and performing rectal douching after homosexual anal intercourse (HR = 3.09, 95%CI = 1.40 - 6.83) were significantly associated with syphilis seroconversion.

CONCLUSIONThe seroconversions of HIV and syphilis during the follow-up visits in this MSM cohort study in Beijing were very serious, and that the associated factors for seroconversions were rectal douching after homosexual anal intercourse and meeting male sex partners in parks, public washrooms or bathhouses.

Adolescent ; Adult ; Antibodies, Bacterial ; blood ; China ; epidemiology ; HIV ; immunology ; HIV Antibodies ; blood ; HIV Infections ; blood ; epidemiology ; HIV Seropositivity ; blood ; epidemiology ; Homosexuality, Male ; Humans ; Incidence ; Male ; Risk Factors ; Sexual Behavior ; Syphilis ; blood ; epidemiology ; Treponema pallidum ; immunology ; Young Adult

To obtain the pure and soluble P51 antigen of HIV-1 strain CN54, we transformed the Escherichia. coli strain BL21 codonplus-RIL with recombinant plasmid pTHioHisA51 which carries a gene encoding the Polymerase (Pol) P51 antigen of HIV-1 CN54 formerly, and induced protein expression by IPTG. We purified the recombinant protein with Chelating Sepharose FF-Ni and DEAE-Sepharose FF column chromatography, then renatured the recombinant protein by dialyzation. Purified protein was identified by Western blotting. We labeled and coated antigen P51 in a dual-antigen sandwich system, and tested it with serum samples from HIV-infected individuals. The results showed that P51 was expressed as inclusion body, and represented about 50% of total cellular protein. After purification and renaturation, the purity of P51 was up to 95%. Western blotting and sandwich ELISA demonstrated that recombinant P51 had good anti-HIV antibody specificity and sensitivity. The results suggested that recombinant HIV-1 P51 can be prepared as diagnostic reagent, and provides valuable support for HIV-1 detection and vaccine research.
Escherichia coli ; genetics ; metabolism ; HIV Antibodies ; blood ; immunology ; HIV Infections ; immunology ; virology ; HIV Reverse Transcriptase ; biosynthesis ; genetics ; immunology ; HIV-1 ; classification ; immunology ; Humans ; Protein Renaturation ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Sensitivity and Specificity

INTRODUCTIONDried blood spot (DBS) collection is an appealing alternative to whole blood or plasma sampling, as it has technical and economic advantages over the latter.

MATERIALS AND METHODSA prospective cross-sectional study was conducted at a Malaysian tertiary referral hospital from November 2009 to March 2010. One hundred and fifty paired specimens of DBS and plasma were analysed by the standard assays for HIV Ag/Ab, HBsAg, anti-HBS and anti-HCV, separately (total 600 paired specimens). DBS sample titres were then compared to the results of plasma testing, which was used as the gold standard.

RESULTSFor the HIV Ag/Ab assay with a cut-off point of 0.35 Relative Light Units (RLUs), the sensitivity and specificity were both 100%. For the HBsAg assay, the sensitivity was 96.5% and the specificity was 97.8%, with a cut-off point of 1.72 RLUs. Sensitivity for the anti-HBs test was 74.2% and the specificity was 86.9%, using a cut-off point of 0.635 RLUs. For the anti-HCV assay, the sensitivity was 97.3% and the specificity was 100%, with a cut-off point of 0.10 RLUs.

CONCLUSIONDBS is an ideal choice to be used as a screening tool for the detection of HIV, Hepatitis B and Hepatitis C virus infections. However, different cut-off values need to be used for the validation of test positivity in DBS samples because the small amount of blood in the DBS specimens leads to lower assay titres.

Cross-Sectional Studies ; Dried Blood Spot Testing ; HIV Antibodies ; blood ; immunology ; HIV Antigens ; blood ; immunology ; HIV Infections ; diagnosis ; Hepacivirus ; isolation & purification ; Hepatitis B ; diagnosis ; Hepatitis B Antibodies ; blood ; immunology ; Hepatitis B Surface Antigens ; blood ; immunology ; Hepatitis C ; diagnosis ; Hepatitis C Antibodies ; blood ; immunology ; Hepatitis C Antigens ; blood ; immunology ; Humans ; Malaysia ; Plasma ; virology ; Prospective Studies ; Referral and Consultation ; Sensitivity and Specificity ; Specimen Handling

It is generally held that HIV, the causative agent of the rampaging global HIV/AIDS pandemic, is incurable and uniformly fatal. Since the discovery and isolation of the virus over two decades ago, global efforts at producing preventive and curative vaccines against it have so far resulted in failure. Working single-handedly with only his family's meagre resources and against the tide of universally accepted dogmas on HIV/AIDS, the author designed, developed and applied new HIV therapeutic and preventive vaccines in Nigeria, and has been using them on willing HIV-infected and normal persons respectively with their informed and written consent since their development. In many cases, the therapeutic vaccine produced rapid improvement not only in the symptoms and signs attributable to HIV infection, but also in various laboratory parameters with a sustained seroconversion to HIV antibody negative status in a number of the patients. In those HIV-infected patients with concomitant hepatitis B (HBV) and/or C (HCV) infection(s), the therapeutic vaccine has resulted in maintained seroconversion to negative (normal) for the HBsAg and HCV antibodies also. No significant adverse or side effect has been observed yet with the use of these vaccines. The vaccines do not cause the production of any detectable levels of stigmatizing anti-HIV antibodies. It is postulated that the vaccines elicit effective but selective cell-mediated cytotoxic immune responses against HIV, HBV and HCV-infected cells.
AIDS Vaccines ; therapeutic use ; Case-Control Studies ; HIV Antibodies ; blood ; HIV Infections ; immunology ; prevention & control ; therapy ; Hepatitis C Antibodies ; blood ; Humans ; Nigeria ; Serologic Tests

OBJECTIVETo study the quantity of anti-R7V in individuals infected with HIV and AIDS patients and its relation with the progression of disease.

METHODSELISA and precipitation and other methods were used to investigate the quantity of anti-R7V in asymptomatic long-term survivors and AIDS patients.

RESULTSPositive rate and quantity of anti-R7V were higher in the HIV active ones and AIDS. It showed that the quantity and positive rate of anti-R7V were rather high in dissolving test.

CONCLUSIONSIt is strong suggestion for anti-R7V to obstruct the replication of virus by interfering the connection between HIV with CCR5 or CXCR4 and so it impossible HIV entering to CD4+ T cells.

Acquired Immunodeficiency Syndrome ; immunology ; Adolescent ; Adult ; Aged ; Child ; Female ; Follow-Up Studies ; HIV Antibodies ; blood ; HIV Infections ; immunology ; HIV Long-Term Survivors ; HIV-1 ; immunology ; Humans ; Male ; Middle Aged ; Receptors, CCR5 ; physiology ; Receptors, CXCR4 ; physiology

BACKGROUNDTo construct replication-deficient recombinant adenovirus expressing wild and codon-modified HIV-1 gp120.

METHODSThe viral codons were changed to the codon usage of highly expressed mammal gene, the resulting modified gp120 gene was synthesized. The wild and modified gp120 genes were cloned into shuttle vector pShuttle-CMV respectively, and then the constructed plasmids containing gp120 gene was cotransformed with the backbone vector pADeasy-1 into E.coli BJ5183. Transfection of the recombinant AdEasy plasmid into 293 cells was performed to obtain recombinant adenoviruses. The mice were immunized with the recombinant adenoviruses. Their immunogenicity was evaluated by testing antibody and CTL levels of immunized mice.

RESULTSTwo strains of recombinant adenovirus expressing wild and codon-modified HIV-1 gp120 were obtained. The protein expressing level of the recombinant adenoviruses containing modified genes was much higher than that containing wild genes. The mice immunized with recombinant adenoviruses elicited HIV-1 specific antibody and CTL response. The rAd-mod gp120 group was better than the rAd-wt gp120 group.

CONCLUSIONReplication-deficient recombinant adenovirus expressing HIV-1 gp120 can elicit HIV-1 specific humoral and cellular response, the codon-modified recombinant virus was more efficient than the native.

AIDS Vaccines ; immunology ; Adenoviridae ; genetics ; Animals ; Codon ; genetics ; Female ; HIV Antibodies ; blood ; HIV Envelope Protein gp120 ; biosynthesis ; genetics ; immunology ; HIV-1 ; immunology ; Mice ; Mice, Inbred BALB C ; Plasmids ; genetics ; Recombination, Genetic ; T-Lymphocytes, Cytotoxic ; immunology