Bagni, Rachel K

Beam, Katie

Bess, Matthew

Burgan, William E

Meade, Allison

Newton, Robert

Wakeham, Katie

Whitby, Denise

English

PubMed

William E Burgan

Katie Beam

Matthew Bess

Allison Meade

Katie Wakeham

Robert Newton

Denise Whitby

Rachel K Bagni

Crossref

POSTER PRESENTATION Open AccessDevelopment of multiplex serological assays todetect oncoviral infectionsWilliam E Burgan1*, Katie Beam1, Matthew Bess1, Allison Meade1, Katie Wakeham2, Robert Newton3,Denise Whitby4, Rachel K Bagni1†From 13th International Conference on Malignancies in AIDS and Other Acquired Immunodeficiencies(ICMAOI)Bethesda, MD, USA. 7-8 November 2011Serological markers of infection (antibodies or antigens) ofviruses that cause cancer are most often detected usingELISA-based methodologies. In many cases, multiple mar-kers of infection must be assessed to determine a finalsero-status. Volume requirements and costs of reagentsfor single analyte ELISAs are high and studies whichinclude multiple viruses can require milliliters of plasma,often not available from archived cohorts. Thus, we soughtto develop a Luminex® bead-based customizable panelinitially including Hepatitis B Virus (HBV), Hepatitis CVirus (HCV), Epstein-Barr Virus (EBV) and often the con-comitant infection, Human Immunodeficiency Virus(HIV) to reduce sample volume requirements, overall costand increase flexibility. Peptides, antigens and antibodieswere sourced from multiple manufacturers and tested forsuitability in this platform. Where necessary, suitablereagents were designed and produced in-house. The HCVassay multiplexes four HCV peptides designed to detectantibodies raised to HCV Core (2), HCV NS4, HCV NS5gene regions. The HBV assay multiplexes a HBV earlyantigen peptide, a recombinant HBV core protein andeither a recombinant HBV surface antigen or an antibodyspecific for HBV surface antigen to assess HBV infection.The EBV assay multiplexes peptides specific to viral capsidantigen, EBNA-1 and early antigen (Cyto-Barr, Zuidhorn,The Netherlands). HIV-1 assay development is ongoingand the list of antigens to be included in the assay has notbeen finalized. These assays can be run singly or with anycombination (multi-plex) of the above listed targets. Eachtarget has been independently validated using samples ofknown molecular and serological status to determinespecificity (false positive versus false negative) and re-evaluated under multiplex conditions to confirm assayperformance. In addition, where possible, samples wereassayed on commercial testing platforms as well as ourmultiplex assay to assess concordance (94-99%). Depen-dent on the panel selected and the expected antibody titersin a particular population, plasma or serum volumes in therange of 10 µL to 125 µL per subject would be required todetermine the HBV, HCV, EBV and/or HIV serostatus ofa subject. This assay platform is inherently flexible and thebenefits include amenability to expansion to include otheroncogenic viruses as well as screening large epidemiologi-cal cohorts or smaller subsets of samples in an economicaland high throughput manner.Author details1Protein Expression Laboratory, Advanced Technology Program, SAIC-Frederick, Frederick, MD, USA. 2Medical Research Council/Uganda VirusResearch Institute Uganda Research Unit on AIDS, Entebbe, Uganda.3Epidemiology and Genetics Unit, Department of Health Sciences, Universityof York, York, UK. 4Viral Oncology Section, AIDS and Cancer Virus Program,SAIC-Frederick, Frederick, MD, USA.Published: 19 April 2012doi:10.1186/1750-9378-7-S1-P30Cite this article as: Burgan et al.: Development of multiplex serologicalassays to detect oncoviral infections. Infectious Agents and Cancer 2012 7(Suppl 1):P30.* Correspondence: burganwe@mail.nih.gov† Contributed equally1Protein Expression Laboratory, Advanced Technology Program, SAIC-Frederick, Frederick, MD, USAFull list of author information is available at the end of the articleBurgan et al. Infectious Agents and Cancer 2012, 7(Suppl 1):P30http://www.infectagentscancer.com/content/7/S1/P30© 2012 Burgan et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative CommonsAttribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction inany medium, provided the original work is properly cited.

Infectious Agents and Cancer

Development of multiplex serological assays to detect oncoviral infections

Springer - Publisher Connector

POSTER PRESENTATION Open Access
Development of multiplex serological assays to
detect oncoviral infections
William E Burgan1*, Katie Beam1, Matthew Bess1, Allison Meade1, Katie Wakeham2, Robert Newton3,
Denise Whitby4, Rachel K Bagni1†
From 13th International Conference on Malignancies in AIDS and Other Acquired Immunodeficiencies
(ICMAOI)
Bethesda, MD, USA. 7-8 November 2011
Serological markers of infection (antibodies or antigens) of
viruses that cause cancer are most often detected using
ELISA-based methodologies. In many cases, multiple mar-
kers of infection must be assessed to determine a final
sero-status. Volume requirements and costs of reagents
for single analyte ELISAs are high and studies which
include multiple viruses can require milliliters of plasma,
often not available from archived cohorts. Thus, we sought
to develop a Luminex® bead-based customizable panel
initially including Hepatitis B Virus (HBV), Hepatitis C
Virus (HCV), Epstein-Barr Virus (EBV) and often the con-
comitant infection, Human Immunodeficiency Virus
(HIV) to reduce sample volume requirements, overall cost
and increase flexibility. Peptides, antigens and antibodies
were sourced from multiple manufacturers and tested for
suitability in this platform. Where necessary, suitable
reagents were designed and produced in-house. The HCV
assay multiplexes four HCV peptides designed to detect
antibodies raised to HCV Core (2), HCV NS4, HCV NS5
gene regions. The HBV assay multiplexes a HBV early
antigen peptide, a recombinant HBV core protein and
either a recombinant HBV surface antigen or an antibody
specific for HBV surface antigen to assess HBV infection.
The EBV assay multiplexes peptides specific to viral capsid
antigen, EBNA-1 and early antigen (Cyto-Barr, Zuidhorn,
The Netherlands). HIV-1 assay development is ongoing
and the list of antigens to be included in the assay has not
been finalized. These assays can be run singly or with any
combination (multi-plex) of the above listed targets. Each
target has been independently validated using samples of
known molecular and serological status to determine
specificity (false positive versus false negative) and re-
evaluated under multiplex conditions to confirm assay
performance. In addition, where possible, samples were
assayed on commercial testing platforms as well as our
multiplex assay to assess concordance (94-99%). Depen-
dent on the panel selected and the expected antibody titers
in a particular population, plasma or serum volumes in the
range of 10 µL to 125 µL per subject would be required to
determine the HBV, HCV, EBV and/or HIV serostatus of
a subject. This assay platform is inherently flexible and the
benefits include amenability to expansion to include other
oncogenic viruses as well as screening large epidemiologi-
cal cohorts or smaller subsets of samples in an economical
and high throughput manner.
Author details
1Protein Expression Laboratory, Advanced Technology Program, SAIC-
Frederick, Frederick, MD, USA. 2Medical Research Council/Uganda Virus
Research Institute Uganda Research Unit on AIDS, Entebbe, Uganda.
3Epidemiology and Genetics Unit, Department of Health Sciences, University
of York, York, UK. 4Viral Oncology Section, AIDS and Cancer Virus Program,
SAIC-Frederick, Frederick, MD, USA.
Published: 19 April 2012
doi:10.1186/1750-9378-7-S1-P30
Cite this article as: Burgan et al.: Development of multiplex serological
assays to detect oncoviral infections. Infectious Agents and Cancer 2012 7
(Suppl 1):P30.
* Correspondence: burganwe@mail.nih.gov
† Contributed equally
1Protein Expression Laboratory, Advanced Technology Program, SAIC-
Frederick, Frederick, MD, USA
Full list of author information is available at the end of the article
Burgan et al. Infectious Agents and Cancer 2012, 7(Suppl 1):P30
http://www.infectagentscancer.com/content/7/S1/P30
© 2012 Burgan et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in
any medium, provided the original work is properly cited.


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Development of multiplex serological assays to detect oncoviral infections

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