The polymerase chain reaction (PCR) was used to detect human cytomegalovirus (HCMV) genome directly without any treatment in urine from 59 renal transplant patients and 49 normal healthy adults. As a virus control, culture supernatant of AD169
strain(ATCC VR-538) were fragments of 133 and 152 bp which encode at major immediate early antigen and late antigen of HCMV.
To enhance the sensitivity, dot blot hybridization ws performed on first and second amplified DNA products using non-radioactive digoxigenin labeled probes. On the basis of four detection methods(e.g.;PCR and gel electrophoresis, PCR and
hybridixation,
two step PCR and gel ellctrophoresis, two step PCR and hyridization), the detection rates in urine samples were compared.
@ES The results were as follows:
@EN 1. The sensitivity was increased to at least 100 fold by two step PCR or hybridization with digoxigenin labeled probe.
2. The PCR and hybridization method was the best among four detection methods because of relatively high detection rate and low nonspecific reaction.
3. By using the PCR and hybridization method, the detection rate in urine freom kidney trans plant patients was 72% using P1/P3 primers and 41% using P4/P5 primers. In contrast, the detection rate in urine from healthy adults was 14% USING P1/P3
primers
and 0% using P4/P5 primers.
This suggest that the PCR and hybridization is very sensitve and can be used for rapid detection of HCMV infections.
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