Determination of plasma viral load in HIV-1 infection by quantitative competitive polymerase chain reaction

Piatak, Michael; Saag, Michael S.; Yang, Limei; Clark, Steven J.; Kappes, John C.; Luk, Ka‐Cheung; Hahn, Beatrice H.; Shaw, George M.; Lifson, Jeffrey D.

doi:10.1097/00002030-199311002-00014

Cited by 55 publications

(29 citation statements)

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“…In humans coinfected with HIV-1 and M. tuberculosis, the magnitude of the virus-mycobacterium coinfection may not be as great as that seen in the macaques coinfected with SIV mac and BCG. The levels of plasma viral RNA in chronically HIV-1-infected humans are usually not as high as those in the SIV mac -infected macaques evaluated in this study (26). Moreover, mycobacterial loads may also be low in humans during a newly transmitted M. tuberculosis coinfection or during reactivation of a latent M. tuberculosis infection, given the clinical difficulty in identifying mycobacteria from specimens collected from patients (4,20).…”

Section: Discussionmentioning

confidence: 73%

Induction of an AIDS Virus-Related Tuberculosis-Like Disease in Macaques: a Model of Simian Immunodeficiency Virus- Mycobacterium Coinfection

Shen¹,

Zhou²,

Chalifoux

et al. 2002

Infect Immun

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Section: Discussionmentioning

confidence: 73%

Induction of an AIDS Virus-Related Tuberculosis-Like Disease in Macaques: a Model of Simian Immunodeficiency Virus- Mycobacterium Coinfection

Shen¹,

Zhou²,

Chalifoux

et al. 2002

Infect Immun

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“…Plasmid pQP1, which contains a 1,420-bp fragment of HIV-1 HXB2 gag (SacI to BglII) downstream of the T7 promoter, was linearized with EcoRI, purified with a PCR purification kit (Qiagen, Valencia, Calif.), and transcribed with T7 RNA polymerase by using the RiboMax Express Large Scale RNA production system (Promega, Madison, Wis.) (24,25). The template DNA was degraded with 5 U of RNase-free DNase, and the RNA transcripts were purified twice with an RNeasy kit (Qiagen).…”

Section: Methodsmentioning

confidence: 99%

New Real-Time Reverse Transcriptase-Initiated PCR Assay with Single-Copy Sensitivity for Human Immunodeficiency Virus Type 1 RNA in Plasma

et al. 2003

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More sensitive assays for human immunodeficiency virus type 1 (HIV-1) RNA are needed to detect, quantify, and characterize persistent viremia in patients who are receiving antiretroviral therapy and whose plasma HIV-1 RNA levels are suppressed to less than 50 to 75 copies/ml. We therefore developed an internally controlled real-time reverse transcriptase-initiated PCR assay that quantifies HIV-1 RNA concentrations down to 1 copy per ml of plasma. This assay with single-copy sensitivity (the single-copy assay) generates a reproducible linear regression plot of input copy number versus threshold cycle by using HIV-1 RNA transcripts at copy numbers ranging from 1 to 10 6 per reaction mixture. The single-copy assay was compared to the ultrasensitive AMPLICOR HIV-1 MONITOR assay and a more sensitive modification of the ultrasensitive assay by repeatedly testing a low-copy-number panel containing 200 to 0.781 copies of HIV-1 RNA per ml of plasma. This comparison showed that the single-copy assay had a greater sensitivity than the other assays and was the only assay that detected HIV-1 RNA at levels as low as 0.781 copies/ml. Testing of plasma samples from 15 patients who were receiving antiretroviral therapy and who had <75 HIV-1 RNA copies/ml revealed persistent viremia in all 15 patients, with HIV-1 RNA levels ranging from 1 to 32 copies/ml (median, 13 copies/ml). The greater sensitivity of the single-copy assay should allow better characterization of persistent viremia in patients who are receiving antiretroviral therapy and whose HIV-1 RNA levels are suppressed to below the detection limits of present assays.

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“…The HIV RNA transcripts were quantified by measuring the specific absorbance at 260 nm, diluted to 10 6 copies/l, divided into aliquots, and stored at Ϫ80°C. The aliquots of HIV-1 RNA transcripts were diluted to generate standards ranging from 10 6 to 0.3 copies/10 l to produce a standard curve for each single-copy assay as described previously (58,59). Threshold cycle values were plotted as a function of the input transcript copy number, and a standard curve was generated by linear regression using ABI 7900 sequence detection software (Applied Biosystems) (52).…”

Section: Methodsmentioning

confidence: 99%

Increased In Vivo Activation of Microglia and Astrocytes in the Brains of Mice Transgenic for an Infectious R5 Human Immunodeficiency Virus Type 1 Provirus and for CD4-Specific Expression of Human Cyclin T1 in Response to Stimulation by Lipopolysaccharides

Sun

Zheng²,

Zhao

et al. 2008

J Virol

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Inflammatory mediators and viral products produced by human immunodeficiency virus (HIV)-infected microglia and astrocytes perturb the function and viability of adjacent uninfected neuronal and glial cells and contribute to the pathogenesis of HIV-associated neurocognitive disorders (HAND). In vivo exposure to lipopolysaccharide (LPS) activates parenchymal microglia and astrocytes and induces cytokine and chemokine production in the brain. HIV-infected individuals display increased circulating LPS levels due to microbial translocation across a compromised mucosa barrier. We hypothesized that HIV-infected microglia and astrocytes display increased sensitivity to the proinflammatory effects of LPS, and this combines with the increased levels of systemic LPS in HIV-infected individuals to contribute to the development of HAND. To examine this possibility, we determined the in vivo responsiveness of HIV-infected microglia and astrocytes to LPS using our mouse model, JR-CSF/human cyclin T1 (JR-CSF/hu-cycT1) mice, which are transgenic for both an integrated full-length infectious HIV type 1 (HIV-1) provirus derived from the primary R5-tropic clinical isolate HIV-1 JR-CSF regulated by the endogenous HIV-1 long terminal repeat and the hu-cycT1 gene under the control of a CD4 promoter. In the current report, we demonstrated that in vivo-administered LPS more potently activated JR-CSF/hu-cycT1 mouse microglia and astrocytes and induced a significantly higher degree of monocyte chemoattractant protein production by JR-CSF/hu-cycT1 astrocytes compared to that of the in vivo LPS response of control littermate mouse microglia and astrocytes. These results indicate that HIV infection increases the sensitivity of microglia and astrocytes to inflammatory stimulation and support the use of these mice as a model to investigate various aspects of the in vivo mechanism of HIV-induced neuronal dysfunction.

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Determination of plasma viral load in HIV-1 infection by quantitative competitive polymerase chain reaction

Cited by 55 publications

References 0 publications

Induction of an AIDS Virus-Related Tuberculosis-Like Disease in Macaques: a Model of Simian Immunodeficiency Virus- Mycobacterium Coinfection

Induction of an AIDS Virus-Related Tuberculosis-Like Disease in Macaques: a Model of Simian Immunodeficiency Virus- Mycobacterium Coinfection

New Real-Time Reverse Transcriptase-Initiated PCR Assay with Single-Copy Sensitivity for Human Immunodeficiency Virus Type 1 RNA in Plasma

Increased In Vivo Activation of Microglia and Astrocytes in the Brains of Mice Transgenic for an Infectious R5 Human Immunodeficiency Virus Type 1 Provirus and for CD4-Specific Expression of Human Cyclin T1 in Response to Stimulation by Lipopolysaccharides

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