Biomedical subjects

H Jayakar

Publications and source records attributed to H Jayakar.

3 recordsLinked to original sources

PubMed2000

Experience with PCR screening.

The COBAS AmpliScreen Hepatitis C (HCV) Test, Version 2.0, which is designed for screening pools composed of samples from individual units of blood or plasma, employs a MultiPrep sample processing procedure that simultaneously extracts and concentrates HCV, HIV-1 and Hepatitis B virus particles from plasma. An HCV Internal Control (IC) RNA serves as an extraction and amplification control for each independently processed sample. Processed samples are amplified by RT-PCR using HCV-specific complementary primers and detected by hybridization of the amplified products to HCV- and IC-specific oligonucleotide probes. The analytical sensitivity of the test is 25 International Units (IU) of HCV per mL of pooled plasma; all HCV genotypes are detected with similar efficiency. The test detected HCV RNA 23 to 32 days prior to anti-HCV antibody seroconversion for four of the five seroconversion panels tested. The test had sufficient sensitivity to reproducibly detect a single infected unit containing 2.4 x 10(3) copies of HCV per mL in a pool with 23 uninfected units. COBAS AmpliScreen tests for HIV-1 and HBV now being validated by Roche Molecular Systems also incorporate the MultiPrep specimen processing method, thereby making it possible to use a single processed specimen to screen for all three viruses.

Genotype↗

PubMed1999

Simultaneous extraction of hepatitis C virus (HCV), hepatitis B virus, and HIV-1 from plasma and detection of HCV RNA by a reverse transcriptase-polymerase chain reaction assay designed for screening pooled units of donated blood.

BACKGROUND: Testing for viral nucleic acids should reduce the residual risk of transmitting viral infections by transfusion of blood components. The AmpliScreen Hepatitis C (HCV) Test, Version 2.0, was designed for screening pools composed of samples from individual units of blood or plasma. STUDY DESIGN AND METHODS: An ultracentrifugation step during sample processing simultaneously extracts and concentrates HCV, HIV type 1, and hepatitis B virus particles from plasma. An HCV internal control RNA serves as an extraction and amplification control for each processed sample. Processed samples are amplified by reverse transcriptase polymerase chain reaction and detected by hybridization of the amplified products to HCV- and internal-control-specific oligonucleotide probes. Plasma samples containing known quantities of HCV were used to evaluate analytical sensitivity and precision. RESULTS: The analytical sensitivity of the test was 25 IU of HCV per mL of pooled plasma; all HCV genotypes were detected with similar efficiency. The test did not react with other blood-borne viruses. The within-run and total coefficients of variation were 1.3-13.0 percent and 1.7-22.0 percent, respectively, with low copy samples producing the more variable results. The test performed well using plasma collected in either EDTA, ACD, or PPT Vacutainer tubes. Plasma samples containing elevated levels of hemoglobin, albumin, triglycerides, or bilirubin did not interfere with the test. The test detected HCV RNA 23 to 32 days prior to seroconversion for four of the five seroconversion panels tested. CONCLUSION: The AmpliScreen HCV Test, Version 2.0 provides a reproducible and specific method for screening pooled blood units for HCV. Theoretically, this test has sufficient sensitivity to detect a single infected unit containing 2.4 x 10(3) IU of HCV per mL in a pool with 95 uninfected units and should reduce the window period by at least 20 to 30 days.

Blood↗

PubMed1998

Ultrasensitive reverse transcription-PCR assay for quantitation of human immunodeficiency virus type 1 RNA in plasma.

With the recent introduction of combination therapy, human immunodeficiency virus type 1 (HIV-1) RNA levels in plasma have been dramatically reduced, frequently to below the limit of quantitation (400 copies/ml of plasma) of the AMPLICOR HIV-1 MONITOR Test (Roche Diagnostic Systems). To achieve enhanced sensitivity of the AMPLICOR HIV-1 MONITOR Test, a modified specimen preparation procedure that allows input of RNA from 10-fold more plasma per amplification reaction was developed. This "ultrasensitive" method allows the accurate quantitation of plasma HIV-1 RNA levels as low as 50 copies/ml. A precision study yielded average within-run and between-run coefficients of variation (CV) of 24.8 and 9.6%, respectively. A multicenter reproducibility study demonstrated that the laboratory-to-laboratory reproducibility of this assay is good, with an average CV of 32%. The linear range of this test is between 50 and 50,000 copies/ml of plasma. RNA concentrations measured by the ultrasensitive and standard HIV-1 MONITOR tests exhibited good agreement within the shared linear range of the two methods. The two measurements were within a factor of 2 for 91% of the specimens tested, with the concentration measured by the ultrasensitive method being only slightly lower (median, 22% lower). Preliminary studies suggest that this assay will prove to be useful for predicting the stability of viral suppression in patients whose RNA levels drop below 400 copies/ml in response to highly active antiretroviral therapy.

Acquired Immunodeficiency Syndrome↗