Development of a rapid diagnostic test for enterovirus-71 using an isothermal amplification based assay
An in-house nucleic acid sequence based amplification (NASBA) assay protocol for the detection of EV-A71 from both virus isolates and clinical specimens was developed in this study. A primer set, targeting the VP1 gene region, was designed and the sensitivity and specificity was tested using conv...
Saved in:
| 主要作者: | |
|---|---|
| 格式: | Thesis |
| 语言: | English |
| 出版: |
2015
|
| 主题: | |
| 在线阅读: | http://ir.unimas.my/id/eprint/10880/1/Ratna%20Sri%20Dewi%20ft.pdf |
| 标签: |
添加标签
没有标签, 成为第一个标记此记录!
|
| 总结: | An in-house nucleic acid sequence based amplification (NASBA) assay protocol for
the detection of EV-A71 from both virus isolates and clinical specimens was developed in
this study. A primer set, targeting the VP1 gene region, was designed and the sensitivity
and specificity was tested using conventional RT-PCR and subsequently used in a NASBA
assay. The detection of EV-A71 NASBA amplicons was successfully achieved using
ethidium bromide-stained RNA agarose gel electrophoresis. The in-house EV-A71 NASBA
protocol detected a panel of 54 of 60 EV-A71 virus isolates from different genogroups and
subgenogroups and no cross-reactivity was observed against other EV serotypes. The
NASBA assay was additionally evaluated on a panel of, 41 clinical specimens of various
sample types from HFMD cases in Sarawak. Twenty-one (95.5%) of 22 clinical specimens
that were confirmed to be EV-A71 positive by RT-PCR and virus isolation were
successfully detected by the NASBA assay. The other 19 clinical specimens that were
concordantly EV-A71 negative by both RT-PCR and virus isolation, were not detected by
the EV-A71 NASBA assay. Verification of the EV-A71 NASBA amplicons from virus
isolates and clinical specimens were successfully achieved by sequencing of the amplified
region. In conclusion, the NASBA assay developed in this study provides a useful
alternative for the screening and detection of EV-A71. |
|---|