Performance characteristics of the COBAS AmpliScreen HIV‐1 test, version 1.5, an assay designed for screening plasma mini‐pools

Yang, Yanfeng; Lamendola, Misty‐Hope; Mendoza, M. Hermoso de; Xu, Dick; Nguyen, Minh; Yeh, Sheria; Wu, Yiching; Ku, Joanne; Rosenstraus, Maurice; Sun, Rita

doi:10.1046/j.1537-2995.2001.41050643.x

Cited by 23 publications

(21 citation statements)

References 15 publications

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“…The calculation that 9 to 16 RNA copies per reaction can be detected with a probability of 95% indicates that all RT-PCRs are well optimized. These copy numbers are close to the detection limits of commercial PCR assays for hepatitis C virus or human immunodeficiency virus (43).…”

Section: Discussionsupporting

confidence: 67%

Rapid Detection and Quantification of RNA of Ebola and Marburg Viruses, Lassa Virus, Crimean-Congo Hemorrhagic Fever Virus, Rift Valley Fever Virus, Dengue Virus, and Yellow Fever Virus by Real-Time Reverse Transcription-PCR

et al. 2002

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Viral hemorrhagic fevers (VHFs) are acute infections with high case fatality rates. Important VHF agents are Ebola and Marburg viruses (MBGV/EBOV), Lassa virus (LASV), Crimean-Congo hemorrhagic fever virus (CCHFV), Rift Valley fever virus (RVFV), dengue virus (DENV), and yellow fever virus (YFV). VHFs are clinically difficult to diagnose and to distinguish; a rapid and reliable laboratory diagnosis is required in suspected cases. We have established six one-step, real-time reverse transcription-PCR assays for these pathogens based on the Superscript reverse transcriptase-Platinum Taq polymerase enzyme mixture. Novel primers and/or 5-nuclease detection probes were designed for RVFV, DENV, YFV, and CCHFV by using the latest DNA database entries. PCR products were detected in real time on a LightCycler instrument by using 5-nuclease technology (RVFV, DENV, and YFV) or SybrGreen dye intercalation (MBGV/EBOV, LASV, and CCHFV). The inhibitory effect of SybrGreen on reverse transcription was overcome by initial immobilization of the dye in the reaction capillaries. Universal cycling conditions for SybrGreen and 5-nuclease probe detection were established. Thus, up to three assays could be performed in parallel, facilitating rapid testing for several pathogens. All assays were thoroughly optimized and validated in terms of analytical sensitivity by using in vitro-transcribed RNA. The >95% detection limits as determined by probit regression analysis ranged from 1,545 to 2,835 viral genome equivalents/ml of serum (8.6 to 16 RNA copies per assay). The suitability of the assays was exemplified by detection and quantification of viral RNA in serum samples of VHF patients.

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Section: Discussionsupporting

confidence: 67%

Rapid Detection and Quantification of RNA of Ebola and Marburg Viruses, Lassa Virus, Crimean-Congo Hemorrhagic Fever Virus, Rift Valley Fever Virus, Dengue Virus, and Yellow Fever Virus by Real-Time Reverse Transcription-PCR

et al. 2002

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“…Use of plasma also allows more precise definition of the experimental target concentrations than do solid samples, e.g., skin crusts. The sensitivities achieved by our assay are comparable to those of commercial US Food and Drug Administration-cleared PCR assays (20,21 ). The good performance of our test in the international ENIVD quality-assurance study on PCR for orthopox viruses further underlines that our test is technically optimized.…”

Section: Discussionmentioning

confidence: 53%

“…In testing of patient samples, PCR inhibition may occur in a frequency of 0.34 -2.1% of tests (20,21 ). The use of inhibition controls is critical to prevent false-negative results (22 ).…”

Section: Discussionmentioning

confidence: 99%

Rapid Detection and Differentiation of Human Pathogenic Orthopox Viruses by a Fluorescence Resonance Energy Transfer Real-Time PCR Assay

et al. 2004

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Background:The orthopox viruses that are pathogenic for humans include variola major virus (VAR), monkeypox virus (MPV), cowpox virus (CPV), and to a lesser extent, camelpox virus (CML) and vaccinia virus (VAC). PCR is a powerful tool to detect and differentiate orthopox viruses, and real-time PCR has the further advantages of rapid turnaround time, low risk of contamination, capability of strain differentiation, and use of multiplexed probes. Methods: We used real-time PCR with fluorescence resonance energy transfer technology to simultaneously detect and differentiate VAR, MPV, CPV/VAC, and CML. An internal control generated by cloning and mutating the PCR target gene facilitated monitoring of PCR inhibition in each individual test reaction. Results: Strain differentiation results showed little interassay variability (CV, 0.4 -0.6%), and the test was 100-fold more sensitive than virus culture on Vero cells. Low copy numbers of DNA could be detected with >95% probability (235-849 genome copies/mL of plasma). Conclusions: The real-time PCR assay can detect and differentiate human pathogenic orthopox viruses. The use of an internal control qualifies the assay for high sample throughput, as is likely to be needed in situations of suspected acts of biological terrorism, e.g., use of VAR.

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“…The reduction in viral load induced by antiretroviral treatment indicates treatment efficacy and correlates with improved clinical outcome (12,26). Detection and quantification of HIV RNA may also help to diagnose HIV-1 infection in children borne by infected mothers (20) and to screen organ (15) and blood (28) donations.…”

mentioning

confidence: 99%

Evaluation of the Clinical Sensitivities of Three Viral Load Assays with Plasma Samples from a Pediatric Population Predominantly Infected with Human Immunodeficiency Virus Type 1 Subtype G and BG Recombinant Forms

Antunes¹,

Figueiredo²,

Bártolo³

et al. 2003

J Clin Microbiol

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Performance characteristics of the COBAS AmpliScreen HIV‐1 test, version 1.5, an assay designed for screening plasma mini‐pools

Abstract: The COBAS AmpliScreen HIV-1 test, v1.5, has sufficient sensitivity to detect a single infected unit containing 600 copies of HIV-1 per mL in a pool with 23 uninfected units and should reduce the window period between infection and seroconversion by at least 2 to 14 days.

Cited by 23 publications

References 15 publications

Rapid Detection and Quantification of RNA of Ebola and Marburg Viruses, Lassa Virus, Crimean-Congo Hemorrhagic Fever Virus, Rift Valley Fever Virus, Dengue Virus, and Yellow Fever Virus by Real-Time Reverse Transcription-PCR

Rapid Detection and Quantification of RNA of Ebola and Marburg Viruses, Lassa Virus, Crimean-Congo Hemorrhagic Fever Virus, Rift Valley Fever Virus, Dengue Virus, and Yellow Fever Virus by Real-Time Reverse Transcription-PCR

Rapid Detection and Differentiation of Human Pathogenic Orthopox Viruses by a Fluorescence Resonance Energy Transfer Real-Time PCR Assay

Evaluation of the Clinical Sensitivities of Three Viral Load Assays with Plasma Samples from a Pediatric Population Predominantly Infected with Human Immunodeficiency Virus Type 1 Subtype G and BG Recombinant Forms

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