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KMID : 0358319950360010017
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Korean Journal of Urology
1995 Volume.36 No. 1 p.17 ~ p.22
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Studies on Detection of Human Immunodeficiency Virus Using Double Polmerase Chain Reaction
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Abstract
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Background:
@EN Serological methods for screening blood and blood products for the presence of antibodies to human immunodeficiency virus (HIV) are efficient and sensitive. In repeatedly reactive cases confirmational tests such as Western blot are available.
However, direct viral detection may be needed for a patient in seronegative window period and a newborn from a infected mother. In addition, a direct assay for the virus would provide a means to monitor both latent and actively replicating virus
in
patients on therpeutic drugs. However, direct detection of HIV in patient samples is difficult and disappointing even with co-cultivation and the successful recovery rate varies from 10 to 75%.
Polymerase chain reaction (PCR) may provide the answer because it can do in vitro amplification of viral genome integrated into human genome (provirus). However, acutal results of clinical application of conventional PCR do not show favorable
sensitivity especially in samples containing very small amounts of HIV molecule copies.
@ES Purpose:
@EN We comparatively analyzed the sensitivity of signle (primary) PCR and double (secondary) PCR in the detection of HIV to define whether double PCR can overcome the limited sensitivity of signle (primary) PCR and if it can be a clinically
promising
method for detecting HIV.
@ES Materials and Methods:
@EN Ten peripheral blood samples from individuals who had antibodies to human immunodeficiency virus were prepared and centrifuged in Ficoll-Hypaque to isolate lymphcytes and monocytes. After DNA extraction from the cell, 35 cycles of primary PCR
was
performed and a part of the PCR product of individual specimen-was electrophoresed to elucidate the results of primary PCR. Secondary PCR with the other part of the individual primary PCR product was followed to compare the efficacies of single
and
double PCR.
@ES Results:
@EN With primary PCR, only one specimen among 10 showed a suspicious corresponding basnd on polyacrylamide gel electrophoresis using ethidium bromide. The results of double PCR presented a striking contrast to those of primary PCR, elucidating
100%
sensitivity without using radioisotope.
@ES Conclusions:
@EN This study suggests that double PCR is a very potent method in detection of human immunodeficiency virus genome incorported in human white blood cells.
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