Objective: To study the diagnostic and epidemiological features of the first two HIV-2 indigenous cases in Hunan province. Methods: Blood samples from two individuals with "HIV antibody indeterminate" and HIV-2 specific band showed by HIV-1/2 western blotting method, were repeatedly collected and detected under HIV 1+2 strip immunoassay and PCR, in Changsha city, Hunan province, through March to November, 2017. An epidemiological survey was carried out at the same time. Results: Our findings showed that the two cases were sex partners, without histories of sexual contact with foreigners and the source of infection was unknown. Results from the HIV 1+2 antibody confirmation test showed that they were "HIV-2 antibody positive" . Through amplifying and sequencing the gag area of HIV-2 and BLAST, the similarity of HIV-2 strains presented as 98%. The results also showed that there were HIV-2 specific fragments in the two cases. Conclusion: HIV-2 indigenous cases had never been reported in China. These cases had brought new challenge on prevention, diagnosis and treatment of HIV/AIDS in China.
Adult ; Blotting, Western/methods* ; China ; HIV Antibodies/isolation & purification* ; HIV Infections/ethnology* ; HIV-2/immunology* ; Humans ; Sexual Behavior ; Sexual Partners ; Surveys and Questionnaires

OBJECTIVETo develop a rapid assay for simultaneous detection of HIV p24 antigen (Ag) and anti-HIV antibody (Ab).

METHODSHIV-1 gp41 antigen and HIV-2 gp36 antigen were expressed by recombinant baculovirus insect system and purified by immunochromatography. p24 monoclonal antibody (mAb) was obtained from p24 hybridoma cell line. Purified antigen and mAb were dot blotted to nitrocellular membrane; 20 nm colloidal gold-anti-human IgG ab and p24 ab complex were used for this test. Previously detected 39 sera specimens were tested in this study to compare with the result of HIV test with commercial HIV test kit.

RESULTS20 mg/L purified gp41 Ag and gp36 Ag were obtained from recombinant baculovirus-insect cell system; 1.5 mg/L p24 mAb was obtained from p24 mAb hybridoma cell line. Compared the test result of 39 sera with commercial HIV test kits, consistency rate was 100%.

CONCLUSIONSThe rapid assay for simultaneous detection of HIV p24 antigen and anti-HIV antibody provides a simple, sensitive and reliable test for HIV diagnosis.

AIDS Serodiagnosis ; Gene Products, env ; biosynthesis ; isolation & purification ; HIV Antibodies ; blood ; HIV Antigens ; biosynthesis ; isolation & purification ; HIV Core Protein p24 ; blood ; HIV Envelope Protein gp41 ; biosynthesis ; isolation & purification ; HIV Infections ; diagnosis ; HIV-1 ; immunology ; HIV-2 ; immunology ; Humans ; Reagent Kits, Diagnostic ; standards ; env Gene Products, Human Immunodeficiency Virus

OBJECTIVETo establish a national reference panel for HIV RNA diagnostic reagents.

METHODSSera from patients with HIV infection and healthy blood donors were collected and tested for HIV and HCV antibodies and HBsAg by using ELISA. The HIV antibody positive samples with ELISA were confirmed with HIV Blot 2.2 (Genelabs). The quantitative samples for HIV RNA were calibrated with the WHO HIV RNA standard. The stability of the panel was evaluated with acceleration method.

RESULTSAfter screening and calibration, 8 negative samples, 8 positive samples, 3 quantitative samples, 6 sensitivity samples and 5 samples for linear analysis were composed of the national reference panel for HIV RNA. The convinced international units (IU) for the quantitative samples were obtained by seven independent calibration and the logarithm of international units for the quantitative samples (b1-b3) were less than x +/- s. The results showed that this panel may stabilize for 4 days at 4 degrees C.

CONCLUSIONA national reference panel for HIV RNA reagents has been established. It may provide the basis for evaluating HIV RNA diagnostic reagents.

Blood Donors ; Calibration ; Drug Stability ; HIV Antibodies ; blood ; HIV Infections ; blood ; virology ; HIV-1 ; genetics ; isolation & purification ; Hepatitis B Surface Antigens ; blood ; Hepatitis C Antibodies ; blood ; Humans ; Indicators and Reagents ; standards ; RNA, Viral ; standards ; Reference Standards ; Reproducibility of Results ; Sensitivity and Specificity

To research piezoelectric immunosensor array for rapid detecting HIV(1+2), piezoelectric immunosensor array matrix was designed. HIV(1+2)C1 antigen was immobilized onto the silver electrodes of quartz-crystal microbalance, which was modified with adsorption and cross-linked method. In the clean air flow and monitoring environment, standard quality control and clinical serum sample were detected. The linear range for the measurement of HIV(1+2) was 0.01-0.2 NCU/ml. The sensitivity, specificity and accuracy of HIV(1+2) piezoelectric protein sensor array were 91.7%, 93.3% and 92.7% respectively.
Biosensing Techniques ; instrumentation ; methods ; Electrodes ; HIV ; isolation & purification ; HIV Antibodies ; blood ; Humans ; Immunoassay ; methods ; Protein Array Analysis ; instrumentation ; methods ; Quartz ; Reproducibility of Results ; Sensitivity and Specificity

To identify human monoclonal HIV-l-neutralizing antibodies from an HIV-1 CRF07BC specific phage display antibody library by cell-based screening. 293T cells were transfected by pCH064. 2-Env plas mid and then used to biopan the phage antibody library. The positive phage clones were screened by cell based ELISA and sequenced for the variable region of heavy (VH) and light (VL) chains. The expressed Fabs were purified by Ni(+2) -NTA column and analyzed by SDS-PAGE. The cell- and gp120 protein-based ELISA as well as flow cytometry were used to measure Fab's binding activity. The neutralizing activity of Fabs was assessed by HIV-1 pseudoviruses. After 4-round biopanning, the binding phages to transfected cells were enriched about 650-folds. A total of 28 positive clones were screened out by cell ELISA and sequence analysis identified 5 different Fabs possessing unique VH and VL (2801, 2837, 2863, 2870 and 2920). Interestingly, these Fabs reacted with the Env-transfected 293T cells but not soluble gp120 proteins, suggesting that they might target conformation-dependent epitopes presenting on viral Env complex. We found that three Fabs (2801, 2863, 2870) exhibited potent neutralizing activity against CRF07_BC isolate CH120. 6 with IC50 of 2.24, 0.89 and 3.09 microg/mL respectively, and that 2801 and 2863 cross-neutral ized the subtype B isolate SF162 at IC50 of 0.69 and 3.52 microg/mL respectively. In conclusion, the HIV-1 Env-transfected 293T cells can be used to efficiently enrich and screen the phage antibody library and isolate human monoclonal HIV-1-neutralizing Fabs that target the Env complex-dependent conformational epitopes. Therefore, our studies provide a powerful platform for exploring the mechanism of HIV-1 neu tralizing response and for designing AIDS vaccines.
Antibodies, Monoclonal ; genetics ; immunology ; Antibodies, Neutralizing ; genetics ; immunology ; Enzyme-Linked Immunosorbent Assay ; methods ; HEK293 Cells ; HIV Antibodies ; genetics ; immunology ; HIV Envelope Protein gp120 ; genetics ; immunology ; HIV Infections ; immunology ; virology ; HIV-1 ; genetics ; immunology ; isolation & purification ; Humans ; Immunoglobulin Fab Fragments ; genetics ; immunology ; Neutralization Tests ; Peptide Library ; Transfection

Objective: To identify the influencing factors that leading to nonspecific responses to indeterminate HIV antibody tests, to provide scientific evidence for the differential diagnosis of HIV infection and control strategy. Methods: A case control study was conducted. The samples of HIV antibody indeterminate in confirmed Western blot (WB) tests, but were negative in HIV nucleic acid tests, were collected as HIV antibody indeterminate group from WB results of HIV confirmatory laboratories of Fujian province in 2015-2016. The general population matched group with HIV antibody screening negative samples and WB negative matched group with WB negative samples were selected as the two compared groups by matching gender and age from HIV antibody screening in Fujian province in the same period. Blood concentrations of hepatitis B surface antigen (HBsAg), anti-hepatitis C virus (HCV) antibody, anti-treponema pallidum (TP) antibody, antinuclear antibody (ANA), anti-human T-cell leukemia virus (HTLV) antibody, and alpha-fetoprotein (AFP) were detected by using enzyme-linked immunosorbent assay (ELISA). χ(2) test and multivariate non-conditional logistic regression analysis were performed to identify the influencing factors that leading to nonspecific responses, to indeterminate HIV antibody tests. Results: A total of 13 WB band patterns were observed in 110 HIV antibody indeterminate samples, in which a single p24 band (58.18%, 64/110), a single gp160 band (17.27%, 19/110) and a single p17 band (7.27%, 8/110) were the three most common patterns. The positive rate of anti-TP antibody was significantly higher in HIV antibody indeterminate samples than general population control group and WB negative control group (10.91%, 12/110 vs. 1.77%, 4/226 and 3.64%, 4/110), compared with two control groups (χ(2)=13.627 and 4.314, P<0.05). The positive rate of AFP was significantly higher in HIV antibody indeterminate samples than general population control group (18.18%, 20/110 vs. 0.44%, 1/226, χ(2)=39.736, P<0.05), the different was not significant compared with WB negative control group (18.18%, 20/110 vs. 23.64%, 26/110, χ(2)=0.990, P>0.05) While no significant differences were found between HIV antibody indeterminate group and two control groups in terms of the positive rates of ANA, HBsAg, anti-HCV antibody or anti-HTLV antibody. Conclusions: The influencing factors that leading to nonspecific responses to indeterminate HIV antibody tests appeared complicate, and the anti-TP antibody positivity might be an influencing factor responsible for nonspecific responses to indeterminate HIV antibody tests.
Adult ; Blotting, Western/methods* ; Case-Control Studies ; China/epidemiology* ; Enzyme-Linked Immunosorbent Assay/methods* ; HIV Antibodies/isolation & purification* ; HIV Infections/epidemiology* ; HIV-1/immunology* ; Humans ; Middle Aged ; Predictive Value of Tests ; Sensitivity and Specificity

OBJECTIVETo investigate the blood quality of blood banks.

METHODSHBsAg, Anti-HCV, Anti-HIV were detected by enzyme linked immunoadsorbent assay (ELISA). Treponema pallidum was detected by Trust or rapid plasma regain card test (RPR) before August 2001 and ELISA was used afterwards. Alanine aminotransferase (ALT) was detected by Reitman-Frankel before December 1999 and by continuous monitoring assay afterwards. Results were compared between (1) sampling and spot check, (2) different determent methods, (3) two kinds of reagents with different antigen coating.

RESULTSSeventy-two (8.28 per thousand ) of the 8,699 plasma samples were found unqualified with levels of anti-HCV, ALT, HBsAg, Treponema pallidum and anti-HIV 24 (2.76 per thousand ), 19 (2.18 per thousand ), 16 (1.84 per thousand ), 7 (0.80 per thousand ), 6 (0.69 per thousand ), respectively. There were significant differences between different determent methods and different reagents, but only ALT showed significant difference in the sampling and spot check.

CONCLUSIONThe unqualified samples were associated with testing methods, quality of reagents as well as ability and responsibility of the staff.

Alanine Transaminase ; blood ; Blood Banks ; standards ; Blood Donors ; Blood-Borne Pathogens ; isolation & purification ; China ; D-Alanine Transaminase ; Enzyme-Linked Immunosorbent Assay ; HIV Antibodies ; blood ; HIV Seropositivity ; Hepatitis B Surface Antigens ; blood ; Hepatitis C Antibodies ; blood ; Humans ; Quality Control

OBJECTIVETo construct and express anti-human RBC and HIVgp160 fusion protein for rapid detection of antibody to HIV.

METHODSThe gene of the anti human RBC ScFv and HIV antigen were constructed together into expression vector. The fusion protein was expressed in E. coli.

RESULTSThe fusion protein was proved to be able to bind both anti-RBC and HIVgp160. It could cause agglutination of human RBC when HIVgp160 was present.

CONCLUSIONThe fusion protein has the potential in rapid detection of HIV.

Antibodies, Monoclonal ; immunology ; isolation & purification ; Autoantibodies ; immunology ; isolation & purification ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Erythrocytes ; immunology ; Gene Expression ; Genetic Vectors ; genetics ; HIV Antibodies ; blood ; immunology ; HIV Envelope Protein gp160 ; genetics ; immunology ; metabolism ; HIV Seropositivity ; blood ; Hemagglutination Tests ; methods ; Humans ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism

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