A novel immunization strategy to induce strong humoral responses against HIV-1 using combined DNA, recombinant vaccinia virus and protein vaccines / 病毒学报

To optimize the immunization strategy against HIV-1, a DNA vaccine was combined with a recombinant vaccinia virus (rTV) vaccine and a protein vaccine. Immune responses against HIV-1 were detected in 30 female guinea pigs divided into six groups. Three groups of guinea pigs were primed with HIV-1 DNA vaccine three times, boosted with rTV at week 14, and then boosted with gp140 protein at intervals of 4, 8 or 12 weeks. Simultaneously, the other three groups of animals were primed with rTV vaccine once, and then boosted with gp140 after 4, 8 or 12 weeks. The HIV-1 specific binding antibody and neutralizing antibody, in addition to the relative affinity of these antibodies, were detected at different time points after the final administration of vaccine in each group. The DNA-rTV-gp140 immune regimen induced higher titers and affinity levels of HIV-1 gp120/gp140 antibodies and stronger V1V2-gp70 antibodies than the rTV-gp140 regimen. In the guinea pigs that underwent the DNA-rTV-gp140 regimen, the highest V1V2-gp70 antibody was induced in the 12-week-interval group. However, the avidity of antibodies was improved in the 4-week-interval group. Using the rTV-gp140 immunization strategy, guinea pigs boosted at 8 or 12 weeks after rTV priming elicited stronger humoral responses than those boosted at 4 weeks after priming. In conclusion, this study shows that the immunization strategy of HIV-1 DNA vaccine priming, followed by rTV and protein vaccine boosting, could strengthen the humoral response against HIV-1. Longer intervals were better to induce V1V2-gp70-specific antibodies, while shorter intervals were more beneficial to enhance the avidity of antibodies.

Identification of Env-specific monoclonal antibodies from Chinese HIV-1 infected person by B cell activation and RT-PCR cloning / 病毒学报

To obtain protective human monoclonal antibody from HIV-1 infected person, we adapted a technology for isolating antigen specific monoclonal antibody from human memory B cells through in vitro B cell activation coupled with RT-PCT and expression cloning. Human B cells were purified by negative sorting from PBMCs of HIV-1 infected individuals and memory B cells were further enriched using anti-CD27 microbeads. Two hundred memory B cells per well were cultured in 96-well round-bottom plates Env-specific antibodies in supernatants were with feeder cells in medium containing EBV and CpG. screened by ELISA after 1-2 weeks' culture. Cells from positive wells of Env-specific antibody were harvested and total RNA was isolated. Human VH and Vkappa or Vlambda genes were amplified by RT-PCR and cloned into IgG1 and kappa or lambda expressing vectors. Functional VH and Vkappa or Vlambda were identified by cotransfecting 293T cells with individual heavy chain and light chain clones followed by analysis of culture supernatants by ELISA for Env-specific antibodies. Finally, corresponding mAb was produced by transient transfection of 293T cells with the identified VH and Vkappa/lambda pair and purified by protein A affinity chromatography. Purified monocolonal antibodies were used for HIV-1 specific antibody-dependent cell-mediated cytotoxicity (ADCC) and neutralizing activity assay. Four monocolonal Env-specific antibodies were isolated from one HIV-1 subtype B' infected individual. Two of them showed strong ADCC activity and one showed weak neutralizing activity against HIV-1. Its further studies on their application in therapeutic or prophylactic vaccines against HIV-1 should be grounded.

Genetic architecture of HIV-1 genes circulating in north India & their functional implications.

This review presents data on genetic and functional analysis of some of the HIV-1 genes derived from HIV-1 infected individuals from north India (Delhi, Punjab and Chandigarh). We found evidence of novel B/C recombinants in HIV-1 LTR region showing relatedness to China/Mynmar with 3 copies of Nfκb sites; B/C/D mosaic genomes for HIV-1 Vpr and novel B/C Tat. We reported appearance of a complex recombinant form CRF_02AG of HIV-1 envelope sequences which is predominantly found in Central/Western Africa. Also one Indian HIV-1 envelope subtype C sequence suggested exclusive CXCR4 co-receptor usage. This extensive recombination, which is observed in about 10 per cent HIV-1 infected individuals in the Vpr genes, resulted in remarkably altered functions when compared with prototype subtype B Vpr. The Vpu C was found to be more potent in causing apoptosis when compared with Vpu B when analyzed for subG1 DNA content. The functional implications of these changes as well as in other genes of HIV-1 are discussed in detail with possible implications for subtype-specific pathogenesis highlighted.

Evaluación de la inmovilización de un péptido de gp36 de VIH-2 en membranas de nitrocelulosa / Evaluation of the immobilization of one HIV-2 gp36 peptide in nitrocellulose membrane

Introducción: la inmovilización de antígenos a soportes sólidos se utiliza para el desarrollo de diversos inmunoensayos. Una de las primeras tecnologías desarrolladas fue la adsorción de proteínas por aplicación directa sobre la nitrocelulosa. Objetivo: normalizar la inmovilización de un péptido sintético de la proteína de transmembrana gp36 del VIH-2, a un soporte de nitrocelulosa para fines diagnósticos y evaluar los parámetros de desempeño en un grupo de muestras de sueros con reactividad de interés conocida. Métodos: el péptido se inmovilizó de forma libre, conjugado a la albúmina de suero bovina (BSA) y a la hemocianina de lapa marina (KLH) como proteínas portadoras. Se analizaron los parámetros de inmovilización y se determinó la variante óptima. Con la variante escogida se evaluó la sensibilidad y especificidad diagnóstica frente a paneles de referencia del Laboratorio de Investigaciones del SIDA. La especificidad analítica se evaluó con muestras reactivas a VIH-1 y HTLV-I. Resultados: el análisis de las variantes de péptido inmovilizadas a las membranas de nitrocelulosa, demostró que el péptido gp36-BSA, fue el que logró la mayor diferenciación entre muestras positivas y negativas. Se obtuvo 100 por ciento de sensibilidad y 95,2 por ciento de especificidad diagnóstica, así como 100 por ciento de especificidad analítica. Conclusiones: el péptido gp36-BSA inmovilizado en membranas de nitrocelulosa es eficaz en el diagnóstico serológico del VIH-2, lo cual permitirá considerarlo para su empleo con fines diagnósticos en sistemas que utilicen como fase sólida la nitrocelulosa.

Introduction: antigen immobilization in solid supports is used for the development of several immunoassays. One of the first technologies developed was the protein adsorption by direct application to nitrocellulose. Objective: to standardize the immobilization of a synthetic peptide of the HIV-2 transmembrane protein gp36 to nitrocellulose support for diagnostic purposes and to evaluate the performance parameters in a group of serum samples with recognized interesting reactivity. Methods: the peptide was freely immobilized, conjugated to bovine serum albumin (BSA) and to keyhole limpet hemocyanin (KLH) as carrier proteins. Immobilization parameters were analyzed and then, the optimal immobilization alternative was determined. Using the chosen variant, the diagnostic sensitivity and specificity against reference panels of the AIDS Research Laboratory were evaluated. Analytical specificity was evaluated with reactive samples to HIV-1 and HTLV-I. Results: the analysis of the immobilized peptide variants to nitrocellulose membranes showed that the gp36 peptide-BSA was the one that succeeded in setting the greatest differentiation between positive and negative samples. There were observed 100 percent sensitivity, 95.2 percent diagnostic specificity and 100 percent analytical specificity. Conclusions: the gp36-BSA peptide immobilized on nitrocellulose membranes showed efficacy for the serological diagnosis of HIV-2, which will allow considering this peptide for diagnostic uses in systems with nitrocellulose based solid phase.

Efficiency of Recombinant Bacille Calmette-Guerin in Inducing Humoral and Cell Mediated Immunities against Human Immunodeficiency Virus Type 1 Third Variable Domain in Immunized Mice

PURPOSE: The third variable (V3) loop of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein has been intensively studied for AIDS vaccine development. Bacille Calmette-Guerin (BCG) is widely used to immunize against tuberculosis and has many advantages as a vaccine vehicle, such as low toxicity, adjuvant potential, low cost, and long-lasting immune-inducing capacity. This work was initiated to investigate the immunogenicity of recombinant BCG (rBCG-mV3) designed to express trimeric HIV-1 V3 loop (mV3) in rBCG-mV3-immunized animals. MATERIALS AND METHODS: HIV-1 V3-concatamer was cloned into pMV261, a BCG-expression vector, and then rBCG-mV3 was constructed by introducing the recombinant plasmid (pMV-V3). The recombinant BCG was examined with regard to its expression of V3-concatamer and the genetic stability in vivo and in vitro. The immune responses induced by recombinant BCG were tested in immunized mice and guinea pigs. RESULTS: The rBCG-mV3 expressed detectable amounts of V3-concatamer when induced by single heat-shock. The recombinant BCG was genetically stable and maintained the introduced mV3 gene for several weeks. V3-specific antibodies were clearly detected 6 weeks after inoculation. The antibody titer rapidly increased after immunization up to 10 weeks, and then maintained for over 4 weeks. IgG2a was prevalent in the V3-specific antiserum. The recombinant BCG was also effective in inducing delayed-type hypersensitivity responses in the immunized guinea pigs. rBCG-immunized mice retained substantial amounts of V3-specific T cells in the spleen, even 5 months after the first immunization. CONCLUSION: Recombinant BCG-mV3 is very efficient in inducing humoral and long-lasting cell-mediated immunity against HIV-1 V3 in the immunized animals.

Efficiency of Recombinant Bacille Calmette-Guerin in Inducing Humoral and Cell Mediated Immunities against Human Immunodeficiency Virus Type 1 Third Variable Domain in Immunized Mice

PURPOSE: The third variable (V3) loop of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein has been intensively studied for AIDS vaccine development. Bacille Calmette-Guerin (BCG) is widely used to immunize against tuberculosis and has many advantages as a vaccine vehicle, such as low toxicity, adjuvant potential, low cost, and long-lasting immune-inducing capacity. This work was initiated to investigate the immunogenicity of recombinant BCG (rBCG-mV3) designed to express trimeric HIV-1 V3 loop (mV3) in rBCG-mV3-immunized animals. MATERIALS AND METHODS: HIV-1 V3-concatamer was cloned into pMV261, a BCG-expression vector, and then rBCG-mV3 was constructed by introducing the recombinant plasmid (pMV-V3). The recombinant BCG was examined with regard to its expression of V3-concatamer and the genetic stability in vivo and in vitro. The immune responses induced by recombinant BCG were tested in immunized mice and guinea pigs. RESULTS: The rBCG-mV3 expressed detectable amounts of V3-concatamer when induced by single heat-shock. The recombinant BCG was genetically stable and maintained the introduced mV3 gene for several weeks. V3-specific antibodies were clearly detected 6 weeks after inoculation. The antibody titer rapidly increased after immunization up to 10 weeks, and then maintained for over 4 weeks. IgG2a was prevalent in the V3-specific antiserum. The recombinant BCG was also effective in inducing delayed-type hypersensitivity responses in the immunized guinea pigs. rBCG-immunized mice retained substantial amounts of V3-specific T cells in the spleen, even 5 months after the first immunization. CONCLUSION: Recombinant BCG-mV3 is very efficient in inducing humoral and long-lasting cell-mediated immunity against HIV-1 V3 in the immunized animals.

Variation of the V3 loop tip motifs and primary drug resistance of HIV-1 strains / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the characteristics of the variation of the V3 loop tip motifs and the drug resistance in the primary treatment patients.</p><p><b>METHODS</b>The partial region of the HIV-1 env and pol gene in 51 samples was amplified by nested polymerase chain reaction (PCR) ,purified products were cloned into the vectors, the obtained were analyzed by MEGA soft wares.</p><p><b>RESULTS</b>The V3 loop tip motifs had four types in our study (GPGR, GPGQ, GPGK, GQGR); the study on the drug resistance in primary treatment patients, showed that there were not major resistance associated with PI, and the resistance were minor mutations in protease gene. In the RT region, there were nine resistance mutants were single NRTIs or NNRTIs.</p><p><b>CONCLUSION</b>The GPGR which was the typical western V3 loop tip motifs attained to 44.44%. This results showed that the percentage of primary drug resistance was still low in our study region, suggesting no need for genotyping detection in blood donor patients before primary therapy.</p>

High genetic diversity in human immunodeficiency virus: type 1 in Jamaica / Elevada diversidad genética del virus de la inmunodeficiencia humana: tipo 1 en Jamaica

The subtypes of the human immunodeficiency virus - type 1 (HIV-1) strains from 54 HIV-1 - infected persons including 44 strains which were typed previously by heteroduplex mobility assay (HMA) were determined by DNA sequencing and phylogenetic analysis. Of 54 HIV- infected persons, 92.5% were infected with HIV-1 subtype B and 7.5% with other HIV-1 subtypes including subtypes D (3.7%), A (1.9%) and J (1.9%). In the phylogenetic analysis, the subtype A virus found in the sample clustered with subtype A reference strains and a circulating recombinant form (CRF) reference strain which originates in Central Africa and is circulating in Cuba indicating a close relationship between these viruses. There was 86% concordance between HMA and DNA sequencing in assigning subtype B viruses. For the non-B subtype viruses, there was less concordance between the two methods (67%). The results confirm the predominance of HIV-1 subtype B strains and the high genetic diversity of HIV-1 strains in circulation in Jamaica. The efficacies and some limitations of the HMA as a method of HIV-1 subtyping also were noted. It is important that the HIV/AIDS epidemic in Jamaica be monitored meticulously for possible expansions in non-B subtypes and the emergence of inter-subtype recombinant forms. We recommend that the more expensive DNA sequencing and phylogenetic analysis, including HIV-1 genotyping for antiretroviral drug resistance testing, be used as an adjunct to the more cost-effective HMA to track the HIV/AIDS epidemic in Jamaica.

Los subtipos de cepas de virus de la inmunodeficiencia humana-tipo-1 de 54 personas infectadas con el VIH-1, que incluyeron 44 cepas previamente clasificadas según su tipo mediante ensayo de movilidad de heterodúplex (HMA), fueron determinados mediante secuenciación de ADN y análisis filogenético. De 54 personas infectados con VIH, 92.5% estaban infectadas con VIH-1 subtipo B y 7.5% con otros subtipos de VIH-1 incluidos los subtipos D (3.7%), A (1.9%), J (1.9%). En el análisis filogenético, el virus de subtipo A hallado en la muestra, se agrupa con las cepas de referencias del subtipo A y una cepa de referencia de forma recombinante circulante (CRF), que tienesu origen en África Central y está circulando en Cuba, lo que indica una estrecha relación entre estos virus. Hubo un 86% de concordancia entre el HMA y la secuenciación del DNA en la asignación de virus de subtipo B. Para los virus de subtipo no B, hubo menos concordancia entre los dos métodos (67%). Los resultados confirman el predominio de las cepas del subtipo B del VIH-1, y la alta diversidad genética de las cepas del VIH-1 en circulación en Jamaica. También se señalaron las eficacias y algunas limitaciones del HMA como método de clasificación del VIH-1 en subtipos. Es importante monitorear meticulosamente la epidemia de VIH/SIDA en Jamaica, a fin de detectar posibles expansiones de subtipos no B y la aparición de formas recombinantes inter-subtipos. Recomendamos que por ser ambos métodos más costosos, tanto la secuenciación de ADN como el análisis filogenético - incluyendo el genotipado del VIH-1 para probar la resistencia antiretroviral del medicamento - sean usados como complementos del HMA, el cual es más costo-efectivo, para seguir de cerca el rastro de la epidemia VIH/SIDA en Jamaica.

Env gene sequencing and HIV-1 subtyping of infected MSM from Beijing / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To comprehend the latest HIV-I epidemic tendency and the character of V3 loop in MSM population of Beijing. METHODS; The C2-V3 regions of the HIV envelop gene were amplified by nest-PCR and sequenced from 11 HIV-l-infected MSM in Beijing in 2007. The subtype and sequences of V3 loop was analyzed. RESULTS There are 4 subtype B strains, 5 CRF AE, 1 CRFO7BC and 1 CRF15-01B strains within all 11 strains. There are five types of central motifs of the 11 samples, in which GPGR and GPGQ are most common.</p><p><b>CONCLUSION</b>Recombination subtype of HIV-1 are spread extensively in MSM population of Beijing.</p>

Genetic analysis of the complete env genes of HIV-1 from paid blood donors in Henan province / 病毒学报

Complete HIV-1 env genes were amplified by nested PCR from uncultured peripheral blood mononuclear cells (PBMCs) DNA of 60 HIV-1 positive paid blood donors in Henan province, and the amplified full-length genes were sequenced. Twenty one full-length env genes were obtained, sequence analysis found that 15 of them had intact open reading frame (ORF). Fourteen sequences conformed to subtype B', their average genetic distance with the international reference sequence RL42 was 4.87% +/- 0.31%. One was subtype B, its genetic distance with the international reference sequence HXB2 was 5.43%. The amino acid sequences of these env genes were deduced according to their nucleotide sequences and extensive analysis and comparison of important structural motifs were performed. The results indicated that there was no drastic alteration in the number and position of potential N-linked glycosylation sites among these 15 sequences. And the residues involved in forming the CD4 binding site were highly conserved. Genotype prediction of coreceptor usage based on V3 sequence and net charge suggested that most samples use CCR5 coreceptor. GPGR motif at the tetrapeptide crown in the V3 loop was most common in these samples and it was detected in 40% sequences. The cleavage site of gp120/gp41 was highly conserved, so Gp160 precursor of all isolates would be efficiently cleaved into the Gp120 and Gp41 subunits. The known neutralizing antibody binding sites for 2G12, IgG1b12, 4E10 and 2F5 were also highly conserved, it is expected that most of these isolates will be sensitive to neutralization by these antibodies. Further study to elucidate the correlation of the env genotype to functionally relevant motifs is necessary and that will aid vaccine and novel drug design.

The impact of the HIV-1 envelope (Env) mutation on its assembly of functional pseudovirus / 病毒学报

To find out whether the mutations of HIV-1 Env have influence on the assembly of pseudovirus and their abilities to infect cells, site-directed mutation (A457D)was performed using cycling mutagenesis and selection of mutants with DpnI. Transformation and plasmid purification technologies were used to obtain mutated env clone. Then both the prototype and the mutant were co-transfected with pSG3(delta(env)) to 293FT cells, respectively. Single-cycle infection assay was employed to analyze the effect of the prototype and the mutant on the ability of functional pseudovirus assembly. The transient expression of both the prototype S12-42-1 and mutant S12-42M were confirmed by Western blot essay. The S/CO value was less than 1 for S12-42-1 and 6.65 for S12-42M, demonstrating the functional pseudovirus was generated only for S12-42M. So mutation on HIV-1 Env has influence on the assembly of pseudovirus and their abilities to infect cells.

Dynamic variation of HIV-1 CRF07_BC env gene in early HIV-1 infection / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To understand the evolution of HIV-1 CRF07_BC envelope, we performed a longitudinal study on two patients during their early HIV-1 infection.</p><p><b>METHODS</b>RNA was extracted from the plasma of the individuals and the C2-C5 fragments of the gp120 gene of HIV-1 were amplified by RT-PCR. Purified DNA segments were inserted into T easy vector and transformed into E. coli Top 10 competent cells. Positive clones were identified by blue-white screening, confirmed by PCR and sequenced by ABI 3700.</p><p><b>RESULTS</b>The samples were collected from the patients every 6 months from seroconversion time. The genetic diversity and divergence in env gene showed consistent increases over time. Our sequence analysis also revealed obvious non-synonymous change in env C1, C3 and V4 regions among these samples.</p><p><b>CONCLUSION</b>The results support the concept that the consistent pattern of viral evolution existed during early phase of HIV-1 infection. C1, C3 and V4 region of env gene may be mainly immunological target during AIDS progression.</p>

Studies on antigencity of human immunodeficiency virus type 1 (HIV-1) external glycoprotein as well as its expression in Pichia pastoris / 生物工程学报

Based on the computer simulation, we analyzed hydrophobicity, potential epitope of recombined subtypes HIV-1 Env protein (851 amino acids) from Guangxi in China. Compared with conservative peptides of other subtypes in env protein, three sequences (469-511aa, 538-674aa, 700-734aa) were selected to recombine into a chimeric gene that codes three conservative epitope peptides with stronger antigencity, and was constructed in the yeast expression plasmid pPICZB. Chimeric proteins were expressed in Pichia pastoris under the induction of methanol, and were analyzed by SDS-PAGE and Westernblot. The results showed that fusion proteins of three-segment antigen were expressed in Pichia pastoris and that specific protein band at the site of 40kD was target protein, which is interacted with HIV-1 serum. The target proteins were purified by metal Ni-sepharose 4B, and were demonstrated to possess good antigenic specificity from the data of ELISA. This chimeric antigen may be used as research and developed into HIV diagnostic reagents.

Subtype and sequence analysis of gag and env genes among HIV-1 strains circulating in Beijing residents during 2006 / 中华流行病学杂志

<p><b>OBJECTIVE</b>To investigate the subtype distribution and the prevalence of sequence characteristics of HIV-1 strains in Beijing residents during 2006 and to analyze the relationship between distribution of HIV-1 subtypes and transmission routines.</p><p><b>METHODS</b>Blood samples from 32 new confirmed HIV-1 infected individuals from Beijing residents in 2006 and separated plasma specimens were collected. RNAs were extracted and the gag and env gene were amplified by RT-PCR and nest-PCR. PCR products were sequenced directly and phylogenetic analyses of gag and env gene were performed using the MEGA2 software.</p><p><b>RESULTS</b>Among 32 HIV-1 plasma samples, 22 gag and 4 env gene fragments were amplified and analyzed. Five HIV-1 subtypes or circulating recombinant forms(CRFs) of HIV-1 including Thai B (2 strains), B (9 strains), C (2 strains), CRF07_BC (5 strains), CRF01 AE (4 strains) were identified being circulated in Beijing. The gene divergences of gag gene inside the subtypes were 6.6%, 4.3%, 6.8%, 4.9% and 3.0% in subtype B, Thai B, C, CRF01_AE and CRF07_BC respectively. Subtypes B were predominant in Beijing, accounted for 40.9% among 22 samples.</p><p><b>CONCLUSION</b>Five HIV-1 subtypes were identified in Beijing and the surveillance of HIV-1 gene variation should be paid more attention to.</p>

Association between sequence variation of Env, Gag genes from the same source and HIV-1 disease progression and host genetic polymorphism / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To understand the relationship between the HIV-1 viral sequence variation and host factors associated with HIV-1 disease progression.</p><p><b>METHODS</b>Env and gag fragments of HIV-1 were amplified with PCR, cloned and sequenced. Bioinformatics was employed to find the genetic variation, N-linked glycosylation, hypermutation etc. Host gene polymorphism was analysed by using restricted fragment length polymorphism (RFLP).</p><p><b>RESULTS</b>Significant difference was found in genetic divergence between Env PCR dominant and clonal sequences (0.1 and 0.06, respectively) in non-treated group, but no significant difference was found in the HAART treated group. V3 GPGQ accounted for the most part in both treated and nontreated groups, rare V3 loop such as GPGH, GQGR and GLGR was found in treated group, V3 substitutions of I/V (position 12) and Y/H (position 21) was associated with the relatively rapid progression (RRP). Glycosylation was significantly higher in RRP than in TP for Env region, GA substitution in RRP was also significantly higher than that in TP group. SDF1-3primeA and CCR2 V64I gene frequency was higher in TP than in RRP, but the difference was not significant.</p><p><b>CONCLUSION</b>Disease progression was associated with V3 AA change, glycosylation and GA substitution in env gene. SDF1-3primeA, CCR2 V64I and CX3CR1 V249I/M280T was not associated with disease progression significantly.</p>

Construction of replication-deficient recombinant adenovirus expressing gag-polDelta and gp140TM genes of human immunodeficiency virus in mice / 中华实验和临床病毒学杂志

<p><b>BACKGROUND</b>Construction of replication-deficient recombinant adenovirus expressing gag-pol and env genes of human immunodeficiency virus (HIV) in mice.</p><p><b>METHODS</b>gag-polDelta and gp140TM genes were cloned into shuttle vector pAdTrack-CMV respectively, and then the plasmids containing gag-polDelta or gp140TM gene were cotransformed with the backbone of adenovirus into E.coli BJ5183. Transfections of the recombinants were performed to obtain recombinant adenoviruses. Its immunogenicity was evaluated by testing antibody levels of mice primed with DNA vaccines and boosted with recombinant adenoviruses.</p><p><b>RESULTS</b>The replication-deficient recombinant adenovirus could express Gp140TM, Gag P55 and P24 proteins correctly. The mice primed with DNA vaccines and boosted with recombinant adenoviruses elicited high titer of HIV-1-specific antibody compared with that inoculated with DNA vaccines only.</p><p><b>CONCLUSION</b>Replication-deficient recombinant adenovirus expressing gag-polDelta and gp140TM can elicit high titer HIV-1-specific antibodies.</p>

Assay for simultaneous detection of HIV p24 antigen and anti-HIV antibody / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To develop a rapid assay for simultaneous detection of HIV p24 antigen (Ag) and anti-HIV antibody (Ab).</p><p><b>METHODS</b>HIV-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.</p><p><b>RESULTS</b>20 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%.</p><p><b>CONCLUSIONS</b>The rapid assay for simultaneous detection of HIV p24 antigen and anti-HIV antibody provides a simple, sensitive and reliable test for HIV diagnosis.</p>