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Ebola Virus (EBOV) Real Time RT-PCR Kit


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Descripción

1. Intended Use
Ebola Virus (EBOV) real time RT-PCR kit is used for the detection of EBOV in serum(non-heparin
anticoagulant), body fluid, or urine sample by using the real time PCR systems.
2. Principle of Real-Time PCR
The principle of the real-time detection is based on the fluorogenic 5’nuclease assay. During the PCR
reaction, the DNA polymerase cleaves the probe at the 5’ end and separates the reporter dye from the
quencher dye only when the probe hybridizes to the target DNA. This cleavage results in the
fluorescent signal generated by the cleaved reporter dye, which is monitored real-time by the PCR
detection system. The PCR cycle at which an increase in the fluorescence signal is detected initially
is proportional to the amount of the specific PCR product. Monitoring the fluorescence intensities in
real-time allows the detection of the accumulating product without having to re-open the reaction
tube after the amplification.
2. Product Description
Ebola is the virus Ebolavirus (EBOV), a viral genus, and the disease Ebola hemorrhagic fever (EHF),
a viral hemorrhagic fever (VHF). The virus is named after the Ebola River Valley in the Democratic
Republic of the Congo (formerly Zaire), which is near the site of the first recognized outbreak in
1976 at a mission hospital run by Flemish nuns. It remained largely obscure until 1989 when several
widely publicized outbreaks occurred among monkeys in the United States.
The virus interferes with the endothelial cells lining the interior surface of blood vessels and with
coagulation. As the blood vessel walls become damaged and destroyed, the platelets are unable to
coagulate, patients succumb to hypovolemic shock. Ebola is transmitted through bodily fluids, while
conjunctiva exposure may also lead to transmission.
EBOV real time RT-PCR kit contains a specific ready-to-use system for the detection of EBOV by
RT-PCR (Reverse Transcription Polymerase Chain Reaction) in the real-time PCR system. The
master contains a Super Mix for the specific amplification of the EBOV RNA. The reaction is done
in one step real time RT-PCR. The first step is a reverse transcription (RT), during which the EBOV
RNA is transcribed into cDNA. Afterwards, a thermostable DNA polymerase is used to amplify the
specific gene fragments by means of PCR (polymerase chain reaction). Fluorescence is emitted and
measured by the real time systems´ optical unit during the PCR. The detection of amplified EBOV
DNA fragment is performed in fluorimeter channel 530nm with the fluorescent quencher BHQ1. In
addition, the kit contains a system to identify possible PCR inhibition by measuring the 560nm
fluorescence of the internal control (IC). An external positive control defined as 1×107copies/ml is
supplied which allow the determination of the gene load. For further information, please refer to
section 9.3 Quantitation.

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