Abstract
Full-length nucleoproteins from Ebola and Marburg viruses were expressed as His-tagged recombinant proteins in Escherichia coli and nucleoprotein-based enzyme-linked immunosorbent assays (ELISAs) were established for the detection of antibodies specific to Ebola and Marburg viruses. The ELISAs were evaluated by testing antisera collected from rabbit immunized with Ebola and Marburg virus nucleoproteins. Although little cross-reactivity of antibodies was observed in anti-Ebola virus nucleoprotein rabbit antisera, the highest reactions to immunoglobulin G (IgG) were uniformly detected against the nucleoprotein antigens of homologous viruses. We further evaluated the ELISA’s ability to detect antibodies to Ebola and Marburg viruses using human sera samples collected from individuals passing through the Guangdong port of entry. With a threshold set at the mean plus three standard deviations of average optical densities of sera tested, the ELISA systems using these two recombinant nucleoproteins have good sensitivity and specificity. These results demonstrate the usefulness of ELISA for diagnostics as well as ecological and serosurvey studies of Ebola and Marburg virus infection.
This is a preview of subscription content, access via your institution.
References
Barrette R W, Metwally S A, Rowland J M, Xu L, Zaki S R, Nichol S T, Rollin P E, Towner J S, Shieh W J, Batten B, Sealy T K, Carrillo C, Moran K E, Bracht A J, Mayr G A, Sirios-Cruz M, Catbagan D P, Lautner E A, Ksiazek T G, White W R, McIntosh M T. 2009. Discovery of swine as a host for the Reston ebolavirus. Science, 325: 204–206.
Bharat T A, Noda T, Riches J D, Kraehling V, Kolesnikova L, Becker S, Kawaoka Y, Briggs J A. 2012. Structural dissection of Ebola virus and its assembly determinants using cryo-electron tomography. Proc Natl Acad Sci USA, 109:4275–4280.
Feldmann H, Jones S, Klenk H D, Schnittler H J. 2003. Ebola virus: from discovery to vaccine. Nat. Rev. Immunol, 3:677–685.
Huang Y, Wei H, Wang Y, Shi Z, Raoul H, Yuan Z. 2012. Rapid detection of filoviruses by real-time TaqMan polymerase chain reaction assays. Virol Sin. 27:273–277.
Ikegami T, Niikura M, Saijo M, Miranda M E, Calaor A B, Hernandez M, Acosta L P, Manalo D L, Kurane I, Yoshikawa Y, Morikawa S. 2003. Antigen capture enzyme-linked immunosorbent assay for specific detection of Reston Ebola virus nucleoprotein. Clin Diagn Lab Immunol, 10:552–557.
Jaax N, Jahrling P, Geisbert T, Geisbert J, Steele K, McKee K, Nagley D, Johnson E, Jaax G, Peters C. 1995. Transmission of Ebolavirus (Zaire strain) to uninfected control monkeys in a biocontainment laboratory. Lancet, 346:1669–1671.
Ksiazek T G, West C P, Rollin P E, Jahrling P B, Peters C J. 1999a. ELISA for the detection of antibodies to Ebola viruses. J Infect Dis, 179:S192–S198.
Ksiazek T G, Rollin P E, Williams A J, Bressler D S, Martin M L, Swanepoel R, Burt F J, Leman P A, Khan A S, Rowe A K, Mukunu R, Sanchez A, Peters C J. 1999b. Clinical virology of Ebola hemorrhagic fever (EHF): virus, virus antigens, and IgG and IgM antibody findings among EHF patients in Kikwit, Democratic Republic of the Congo, 1995. J Infect Dis, 179:S177–S189.
Kuhn J H, Bao Y M, Bavari S, Becker S, Bradfute S, Brister J R, Bukreyev A A, Cai Y, Chandran K, Davey R A, Dolnik O, Dye J M, Enterlein S, Gonzalez J P, Formenty P, Freiberg A N, Hensley L E, Honko A N, Ignatyev G M, Jahrling P B, Johnson K M, Klenk H D, Kobinger G, Lackemeyer M G, Leroy E M, Lever M S, Lofts L L, Mühlberger E, Netesov S V, Olinger G G, Palacios G, Patterson J L, Paweska J T, Pitt L, Radoshitzky S R, Ryabchikova E I, Saphire E O, Shestopalov A M, Smither S J, Sullivan N J, Swanepoel R, Takada A, Towner J S, van der Groen G, Volchkov V E, Wahl-Jensen V, Warren T K, Warfield K L, Weidmann M, Nichol S T. 2013. Virus nomenclature below the species level: a standardized nomenclature for laboratory animal-adapted strains and variants of viruses assigned to the family Filoviridae. Arch Virol, 158:1425–1432.
Leroy E M, Kumulungui B, Pourrut X, Rouquet P, Hassanin A, Yaba P, De’licat A, Paweska J T, Gonzalez J P, Swanepoel R. 2005. Fruit bats as reservoirs of Ebola virus. Nature, 438:575–57.
Lötfering B, Mühlberger E, Tamura T, Klenk H, Becker S. 1999. The Nucleoprotein of Marburg Virus Is Target for Multiple Cellular Kinases. Virology, 255:50–62.
Nakayama E, Yokoyama A, Miyamoto H, Igarashi M, Kishida N, Matsuno K, Marzi A, Feldmann H, Ito K, Saijo M, Takada A. 2010. Enzyme-Linked Immunosorbent Assay for Detection of Filovirus Species-Specific Antibodies. Clin Vaccine Immunol, 17:1723–1728.
Negredo A, Palacios G, Vazquez-Moron S, Gonzalez F, Dopazo H, Molero F, Juste J, Quetglas J, Savji N, de la Cruz M M, Herrera JE, Pizarro M, Hutchison S K, Echevarria J E, Lipkin W I, Tenorio A. 2011. Discovery of an ebolavirus-like filovirus in europe. PLoS Pathog, 7: e1002304.
Niikura M, Ikegami T, Saijo M, Kurane I, Miranda M E, Morikawa S. 2001. Detection of Ebola viral antigen by enzyme-linked immunosorbent assay using a novel monoclonal antibody to nucleoprotein. J Clin Microbiol, 39: 3267–3271.
Niikura M, Ikegami T, Saijo M, Kurata T, Kurane I, Morikawa S. 2003. Analysis of linear B-cell epitopes of the nucleoprotein of ebola virus that distinguish ebola virus subtypes. Clin Diagn Lab Immunol, 10, 83–87.
Ou W, Delisle J, Konduru. K, Bradfute S, Radoshitzky S R, Retterer C, Kota K, Bavari S, Kuhn J H, Jahrling P B, Kaplan G, Wilsona G A. 2011. Development and characterization of rabbit and mouse antibodies against ebolavirus envelope glycoproteins. J Virol Methods, 173:99–109.
Prehaud C, Hellebrand E, Coudrier D, Volchkov V E, Volchkova VA, Feldmann H, Le Guenno B, Bouloy M. 1998. Recombinant Ebola virus nucleoprotein and glycoprotein (Gabon 94 strain) provide new tools for the detection of human infections. J Gen Virol, 79:2565–2572.
Saijo M, Niikura M, Morikawa S, Ksiazek T G, Meyer R F, Peters C J, Kurane I. 2001. Enzyme-linked immunosorbent assays for detection of antibodies to Ebola and Marburg viruses using recombinant nucleoproteins. J Clin Microbiol, 39:1–7.
Saijo M, Niikura M, Ikegami T, Kurane I, Kurata T, Morikawa S. 2006. Laboratory Diagnostic Systems for Ebola and Marburg Hemorrhagic Fevers Developed with Recombinant Proteins. Clin Vaccine Immunol, 13:444–451.
Sanchez A, Kiley M P, Klenk H D, Feldmann H. 1992. Sequence analysis of the Marburg virus nucleoprotein gene: comparison to Ebola virus and other non-segmented negative-strand RNA viruses. J Gen Virol, 73:347–357.
Sanchez A, Geisbert T W, Feldmann H. 2007. In: Filoviridae: Marburg and Ebola viruses. Knipe D M, Howley P M (eds), Fields Virology, 5th ed, Philadelphia: Lippincott Williams and Wilkins, pp 1409–1448.
Towner J S, Amman B R, Sealy T K, Carroll S A, Comer J A, Kemp A, Swanepoel R, Paddock C D, Balinandi S, Khristova M L, Formenty P B, Albarino C G, Miller D M, Reed Z D, Kayiwa J T, Mills J N, Cannon D L, Greer P W, Byaruhanga E, Farnon E C, Atimnedi P, Okware S, Katongole-Mbidde E, Downing R, Tappero J W, Zaki S R, Ksiazek T G, Nicho S T, Rollin P E. 2009. Isolation of genetically diverse Marburg viruses from Egyptian fruit bats. PLoS Pathog, 5:e1000536.
van der Groen G, Kurata T, Mets C. 1983. Modifications to indirect immunofluorescence tests on Lassa, Marburg, and Ebola materials. Lancet, 1:654–655.
Watanabe S, Noda T, Kawaoka Y. 2006. Functional mapping of the nucleoprotein of Ebola virus. J Virol, 80:3743–3751.
WHO: EBOLA RESPONSE ROADMAP UPDATE 26 September 2014. http://apps.who.int/iris/bitstream/10665/135029/1/roadmapupdate26sept14_eng.pdf.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Huang, Y., Zhu, Y., Yang, M. et al. Nucleoprotein-based indirect enzyme-linked immunosorbent assay (indirect ELISA) for detecting antibodies specific to Ebola virus and Marbug virus. Virol. Sin. 29, 372–380 (2014). https://doi.org/10.1007/s12250-014-3512-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12250-014-3512-0
Keywords
- Ebola virus
- Marburg virus
- recombinant nucleoproteins
- indirect ELISA