Developing a Salivary Antibody Multiplex Immunoassay to Measure Human Exposure to Environmental Pathogens.
The etiology and impacts of human exposure to environmental pathogens are of major concern worldwide and, thus, the ability to assess exposure and infections using cost effective, high-throughput approaches would be indispensable. This manuscript describes the development and analysis of a bead-based multiplex immunoassay capable of measuring the presence of antibodies in human saliva to multiple pathogens simultaneously. Saliva is particularly attractive in this application because it is noninvasive, cheaper and easier to collect than serum. Antigens from environmental pathogens were coupled to carboxylated microspheres (beads) and used to measure antibodies in very small volumes of human saliva samples using a bead-based, solution-phase assay. Beads were coupled with antigens from Campylobacter jejuni, Helicobacter pylori, Toxoplasma gondii, noroviruses (G I.1 and G II.4) and hepatitis A virus. To ensure that the antigens were sufficiently coupled to the beads, coupling was confirmed using species-specific, animal-derived primary capture antibodies, followed by incubation with biotinylated anti-species secondary detection antibodies and streptavidin-R-phycoerythrin reporter (SAPE). As a control to measure non-specific binding, one bead set was treated identically to the others except it was not coupled to any antigen. The antigen-coupled and control beads were then incubated with prospectively-collected human saliva samples, measured on a high throughput analyzer based on the principles of flow cytometry, and the presence of antibodies to each antigen was measured in Median Fluorescence Intensity units (MFI). This multiplex immunoassay has a number of advantages, including more data with less sample; reduced costs and labor; and the ability to customize the assay to many targets of interest. Results indicate that the salivary multiplex immunoassay may be capable of identifying previous exposures and infections, which can be especially useful in surveillance studies involving large human populations.
National Exposure Research Laboratory, U.S. Environmental Protection Agency; Augustine.Swinburne@epa.gov.
Swinburne A J
SA
Augustine
National Risk Management Research Laboratory, U.S. Environmental Protection Agency.
Tarsha N
TN
Eason
National Exposure Research Laboratory, U.S. Environmental Protection Agency.
Kaneatra J
KJ
Simmons
Oak Ridge Institute for Science and Education.
Clarissa L
CL
Curioso
National Exposure Research Laboratory, U.S. Environmental Protection Agency.
Shannon M
SM
Griffin
National Exposure Research Laboratory, U.S. Environmental Protection Agency.
Malini K D
MK
Ramudit
Department of Biological Sciences, McMicken College of Arts and Sciences, University of Cincinnati.
Trevor R
TR
Plunkett
D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000276:immunology; D000942:Antigens, Bacterial / Q000032:analysis / Q000276:immunology; D000956:Antigens, Viral / Q000032:analysis / Q000276:immunology; D004781:Environmental Exposure / Q000032:analysis*; D004783:Environmental Microbiology; D016480:Helicobacter pylori / Q000276:immunology; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D007159:Immunologic Tests; D008863:Microspheres; D029322:Norovirus / Q000276:immunology; D012463:Saliva / Q000276:immunology; D014122:Toxoplasma / Q000276:immunology
D000906:Antibodies; D000942:Antigens, Bacterial; D000956:Antigens, Viral
J Clin Pathol. 2000 Apr;53(4):314-7
10823129
Risk Anal. 2004 Feb;24(1):221-36
15028014
Clin Diagn Lab Immunol. 2004 Nov;11(6):1028-34
15539501
J Dent Res. 1987 Feb;66(2):420-4
3305624
Clin Diagn Lab Immunol. 2004 Nov;11(6):1045-53
15539504
Vaccine. 2008 Aug 12;26(34):4403-9
18585420
J Clin Microbiol. 2000 Jun;38(6):2344-7
10835000
J Food Prot. 2011 Mar;74(3):373-9
21375872
J Immunol Methods. 2015 Sep;424:53-63
25985985
J Vet Med Sci. 2000 Nov;62(11):1231-4
11129874
J Med Microbiol. 2003 Jul;52(Pt 7):569-72
12808078
Toxicology. 2004 May 20;198(1-3):229-38
15138046
PLoS Negl Trop Dis. 2014 Jul 31;8(7):e3040
25078404
J Immunol Methods. 2011 Feb 1;364(1-2):83-93
21093445
Malar J. 2011 Apr 29;10:104
21527045
J Water Health. 2003 Sep;1(3):125-31
15384723
J Oral Pathol Med. 1990 Mar;19(3):119-25
2187975
J Immunol Methods. 2006 Feb 20;309(1-2):200-4
16406059
J Virol Methods. 2011 May;173(2):169-74
21295610
Indian J Med Microbiol. 2008 Oct-Dec;26(4):327-32
18974484
J Immunol Methods. 2015 Oct;425:1-9
26070441
PLoS Negl Trop Dis. 2011 May 10;5(5):e1046
21572982
Water Sci Technol. 2004;50(2):23-30
15344769
Clin Diagn Lab Immunol. 1999 Jul;6(4):577-80
10391866
BMJ. 1992 Jul 25;305(6847):207-8
1290500
Epidemiol Infect. 2006 Dec;134(6):1303-12
16650326
J Infect Dis. 2000 Oct;182(4):1260-3
10979930