CDC DENV-1-4 Real-Time RT-PCR Assay

New! FAQ for RT-PCR kit

The CDC Real Time RT-PCR Assay for Dengue Diagnosis

Dengue is one of the most common causes of fever in travelers returning from South East Asia, South Central Asia, and the Caribbean and Latin America. Dengue virus (DENV) infection can be divided into two phases: acute and convalescent. The acute phase begins with the onset of symptoms and lasts approximately 5 days. The convalescent phase begins once the fever subsides and lasts 4 to 7 days.

Patients with dengue usually have symptoms when DENV is present in the blood. Virus levels in the blood are usually higher during the first 5 days of the illness. The CDC Real Time RT-PCR assay detects DENV in a significantly high proportion of cases during this time, very early after the onset of symptoms of dengue. This is important because an early laboratory confirmation of a dengue diagnosis helps physicians managing the case.

The CDC developed this assay using the Applied Biosystems (ABI) 7500 Fast Dx Real-Time PCR Instrument, used for influenza testing. This equipment was distributed by CDC to over 100 US and international public health laboratories, as part of the response to the H1N1 epidemic. CDC adapted their dengue RT-PCR test to take advantage of this widely used testing platform in order to facilitate the use of molecular testing for dengue diagnosis.

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Characteristics of the CDC Real Time RT-PCR Assay

The CDC DENV-1-4 Real-Time RT-PCR Assay is the first nucleic acid diagnostic device for detection and typing of DENV nucleic acid in suspected, symptomatic cases approved by the FDA for use in the United States. This assay detects DENV serotypes 1, 2, 3 or 4 from human serum or plasma collected from human patients with signs and symptoms consistent with dengue infection. The assay is intended to be used as a diagnostic test in patients and is not approved for blood bank screening.

The CDC DENV-1-4 Real-Time RT-PCR Assay includes:

• Oligonucleotide primers and dual-labeled hydrolysis (Taqman®) probes for in vitro qualitative detection of dengue virus serotypes 1, 2, 3 or 4 from serum or plasma collected from human patients with signs and symptoms consistent with dengue (mild or severe).
• Positive control virus mix, which consists of heat-inactivated DENV-1 Haw, DENV-2 NGC, DENV-3 H87, and DENV-4 H241.
• Human Specimen Control (HSC) is a noninfectious cultured human cell material that provides a positive signal in the assay and demonstrates successful recovery of RNA as well as the integrity of the RNA extraction reagent.

The human RNase P RNA (RP) is present in cultured cell material and in most clinical samples and detectable by RT-PCR using the primers and probes provided. The device does not include ancillary reagents.

The CDC DENV-1-4 Real-Time RT-PCR Assay can be run in singleplex (each DENV serotype detected in a separate reaction) or in multiplex (the four DENV serotypes are run in the same reaction). These two formats provide equal sensitivity.

 

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Intended Use of the CDC DENV-1-4 Real Time RT-PCR Assay

The CDC DENV-1-4 Real-Time RT-PCR Assay is intended for use on an Applied Biosystems (ABI) 7500 Fast Dx Real-Time PCR Instrument:

• For the diagnosis of dengue in serum or plasma collected from patients with signs and symptoms consistent with dengue (mild or severe);
• For the identification of dengue virus serotypes 1, 2, 3 or 4 from viral RNA in serum or plasma (sodium citrate) collected from human patients with dengue;
• To provide epidemiologic information for surveillance of circulating dengue viruses.
• Testing of clinical blood specimens (serum or plasma) with the CDC DENV-1-4 Real-Time RT-PCR Assay should not be performed unless the patient meets clinical and/or epidemiologic criteria for testing suspect dengue cases.

The CDC DENV-1-4 Real-Time RT-PCR Assay is not approved for the screening of blood or plasma donors.

Positive results on the CDC DENV-1-4 Real Time RT-PCR Assay indicate current dengue infection. Blood samples from patients suspected of having dengue taken 1-5 days after fever onset should be tested with this device. Negative results obtained with this test do not preclude the diagnosis of dengue and should not be used as the sole basis for treatment or other patient management decisions. If RT-PCR results are negative, anti-DENV IgM testing should be considered (E.g. Inbios DENV Detect IgM Capture ELISA). If the blood sample is taken from a patient 5 or more days after the onset of symptoms, laboratory diagnosis is best made using a test for IgM antibody to DENV.

Assay Distribution

The CDC DENV-1-4 Real Time RT-PCR Assay will be distributed to laboratories with personnel who have training and experience in standardized molecular diagnostic testing procedures and viral diagnosis, and appropriate biosafety equipment and containment.

Please click if your laboratory is in the United States (including Puerto Rico, Guam, American Samoa and affiliated territories).

Information on the CDC DENV-1-4 Real Time RT-PCR Assay for Laboratories in the United States

Materials Provided in the CDC DENV-1-4 RT-PCR Assay

• Box 1: Detection Kit (Primer and Probe Sets)
• Box 2: Positive Control Kit (A mix of heat inactivated DENV-1, -2, -3 and -4 standards)
• Box 3: Human Specimen Extraction Control (HSC)

Click to order the CDC DENV-1-4 Real-Time RT-PCR Assay or for technical support.

Materials Required (But Not Provided)

The following is a list of ancillary reagents that are not supplied with the CDC DENV-1-4 Real-Time RT-PCR Assay. The Invitrogen and Roche products are included in CDC’s reagent qualification program.

*The CDC DENV-1-4 Real-Time RT-PCR Assay test performance requires that only qualified ancillary reagent lots be used with the device in the United States. Any lots not specifically qualified by the CDC-Dengue Branch for use with the CDC DENV-1-4 Real-Time RT-PCR Assay are not valid for use with this device, and may affect device performance.

** Qiagen QIAamp® DSP Viral RNA Mini Kit (Cat # 61904) is produced under Good Manufacturing Practices (GMP). These RNA extraction kits can be used manually or in combination with the QIAcube Instrument (Cat # 9001292).

Other consumables and minor equipment are required as specified in the package insert of the CDC DENV-1-4 Real Time RT-PCR Assay.