Skip to main content

Real-Time and End-Point PCR Diagnostics for Ebola Virus

  • Protocol
  • First Online:
Ebolaviruses

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1628))

Abstract

Reverse transcriptase polymerase chain reaction (RT-PCR)-based techniques allow for highly sensitive and specific detection of RNA viruses. Detection of the amplification products can be achieved using several methods. The following are descriptions of the detection of ebolavirus RNA using end-point RT-PCR (agarose gel visualization of amplification products) and quantitative RT-PCR (Q-RT-PCR), with fluorescent detection using an intercalating dye or detection with the use of 5′ hydrolysis probe assays. All of these techniques can be used to accurately detect the presence of ebolavirus in samples.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Sanchez A, Ksaizek TG, Rollin PE, Miranda MEG, Trappier SG, Khan AS et al (1999) Detection and molecular characterization of ebola viruses causing disease in human and nonhuman primates. J Infect Dis 179(Suppl 1):S164–S169

    Article  CAS  PubMed  Google Scholar 

  2. Towner J, Sealy T, Khristova M, Albariño C, Conlan S, Reeder S et al (2008) Newly discovered ebola virus associated with hemorrhagic fever outbreak in Uganda. PLoS Pathog 4(11):e1000212

    Article  PubMed  PubMed Central  Google Scholar 

  3. Grolla A, Jones SM, Fernando L, Strong JE, Ströher U, Möller P et al (2011) The use of a mobile laboratory unit in support of patient management and epidemiological surveillance during the 2005 Marburg outbreak in Angola. PLoS Negl Trop Dis 5(5):e1183

    Article  PubMed  PubMed Central  Google Scholar 

  4. Towner J, Rollin PE, Bausch DG, Sanchez A, Crary SM, Vincent M et al (2004) Rapid diagnosis of Ebola hemorrhagic fever by reverse transcription-PCR in an outbreak setting and assessment of patient viral load as a predictor of outcome. J Virol 78(8):4330–4341

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Trombley A, Wachter L, Garrison J, Buckley-Beason V, Jarhling J, Hensley L et al (2010) Comprehensive panel of real-time TaqMan polymerase chain reaction assays for detection and absolute quatification of filoviruses, arenaviruses and New World hantaviruses. Am J Trop Med Hyg 82(5):954–960

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Panning M, Laue T, Olschlager S, Eickmann M, Becker S, Raith S et al (2007) Diagnostic reverse-transcription polymerase chain reaction kit for filoviruses based on the strain collections of all European biosafety level 4 laboratories. J Infect Dis 196(Suppl 2):S199–S204

    Article  CAS  PubMed  Google Scholar 

  7. Pinsky BA, Sahoo MK, Sandlund J, Kleman M, Kulkarni M, Grufman P et al (2015) Analytical performance characteristics of the Cepheid GeneXpert Ebola assay for the detection of ebola virus. PLoS One 10(11):e0142216

    Article  PubMed  PubMed Central  Google Scholar 

  8. Dreier J, Störmer M, Kleesiek K (2005) Use of bacteriophage MS2 as an internal control in viral reverse transcription-PCR assays. J Clin Microbiol 43(9):4551–4557

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Boddicker JD, Rota PA, Kreman T, Wangeman A, Lowe L, Hummel KB et al (2007) Real-time reverse transcription PCR assay for detection of mumps virus RNA in clinical specimens. J Clin Microbiol 45(9):2902–2908

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Smither SJ, Weller SA, Phelps A, Eastaugh L, Ngugi S, O’Brien LM et al (2015) Buffer AVL alone does not inactivate ebola virus in a representative clinical sample type. J Clin Microbiol 53(10):3148–3154

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Allen Grolla .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2017 Springer Science+Business Media LLC

About this protocol

Cite this protocol

Grolla, A. (2017). Real-Time and End-Point PCR Diagnostics for Ebola Virus. In: Hoenen, T., Groseth, A. (eds) Ebolaviruses. Methods in Molecular Biology, vol 1628. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7116-9_27

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-7116-9_27

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7115-2

  • Online ISBN: 978-1-4939-7116-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics