CDC DENV-1-4 Real-Time rRT-PCR Multiplex Assay

Key points

  • Nucleic acid amplification tests (NAAT) can detect dengue virus infections during the first 7 days of illness.
  • CDC's DENV-1-4 rRT-PCR assay is a NAAT for detection and typing of dengue viruses.
  • CDC's DENV-1-4 rRT-PCR assay is approved for diagnosis of dengue by the U.S. Food and Drug Administration (FDA) and distributed to laboratories.
Three vials of testing solution sitting on ice with a pipette extracting liquid from the one in the center.

Characteristics of the CDC DENV-1-4 rRT-PCR multiplex assay

The CDC DENV-1-4 rRT-PCR assay is a NAAT for detection and typing of dengue virus in suspected, symptomatic dengue cases. This assay detects dengue virus serotypes 1, 2, 3, or 4 from serum or plasma collected from human patients.

The CDC DENV-1-4 rRT-PCR multiplex assay kit 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 CDC DENV-1-4 rRT-PCR multiplex 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.

Intended use of the CDC

The CDC DENV-1-4 rRT-PCR multiplex assay is intended for use on an Applied Biosystems (ABI) 7500 Fast Dx Real-Time PCR Instrument for the following purposes:

  • Diagnosis of dengue in serum or plasma collected from patients with signs and symptoms consistent with dengue.
  • Identification of DENV 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
  • This assay should be used on specimens collected during the first 7 days after the patient's symptom onset.

Keep in mind‎

This assay should be used on specimens collected during the first 7 days after the patient's symptom onset.

The CDC DENV-1-4 rRT-PCR multiplex assay is not approved for the screening of blood or plasma donors.

Interpretation of results

  • Positive results on the CDC DENV-1-4 rRT-PCR multiplex assay indicate confirmed current dengue infection.
  • Negative results obtained with this test do not rule out the diagnosis of dengue.
    • IgM testing is recommended as part of the testing algorithm and may confirm diagnosis.
  • If the blood sample is taken from a patient after 7 or more days after the onset of symptoms, serologic testing for IgM antibodies to DENV is preferred.
    • IgM antibodies against dengue virus can remain detectable for 3 months or longer after infection.

Assay distribution

The CDC DENV-1-4 rRT-PCR multiplex assay is 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. Requests can be sent to DenguePCROrdering@cdc.gov.

Information for laboratories in the United States

Materials provided in the CDC DENV-1-4 rRT-PCR multiplex 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)

See package insert for the United States here (print only) [PDF – 53 pages]

Kit: rRT-PCR enzyme mastermix options

Reagent: Invitrogen SuperScript™ III Platinum® reactions One-Step Quantitative RT-PCR System (without Rox)*

  • Quantity – 100 reactions
    • Catalog No: 11732-020
  • Quantity – 500 reactions
    • Catalog No: 11732-088

Kit: nucleic acid purification kit options

Reagent: Qiagen QIAamp® DSP Viral RNA Mini Kit and Qiagen QIAcube Instrument**

  • Quantity – 50 Extractions
    • Catalog No: 71904 and 9001292

Reagent: Roche MagNA Pure LC total Nucleic Acid Isolation Kit *

  • Quantity – 192 Extractions
    • Catalog No: 03 038 505 001

*The CDC DENV-1-4 rRT-PCR multiplex 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 rRT-PCR multiplex assay are not valid for use with this device and may affect device performance.

** Qiagen QIAamp® DSP Viral RNA Mini Kit (Cat # 71904) is produced under Good Manufacturing Practices (GMP). These RNA extraction kits can be used manually or in combination with the QIAcube Instrument (Cat # 9001292).

Information on the CDC DENV-1-4 rRT-PCR multiplex assay for international laboratories

Materials provided in the CDC DENV-1-4 rRT-PCR multiplex 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)

See international package insert here (print only) [PDF – 51 pages]

Contact information, ordering, and product support

For technical and product support and to obtain the CDC DENV-1-4 rRT-PCR multiplex assay and a list of qualified ancillary reagents, please contact the CDC Dengue Branch support team directly.

For ordering and technical support send email to: DenguePCROrdering@cdc.gov.

Include the following information in your message:

  • Laboratory name and address
  • Qualified contact person
  • Phone number
  • Email address
  • Shipping address
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Content Source:
National Center for Emerging and Zoonotic Infectious Diseases (NCEZID)