A Novel Multiplexed Enzyme-Linked Immunosorbent Assay for the Detection of IgG Seroreactivity to Cytomegalovirus (CMV) UL144

A Novel Multiplexed Enzyme-Linked Immunosorbent Assay for the Detection of IgG Seroreactivity to Cytomegalovirus (CMV) UL144

Infection with human cytomegalovirus (CMV) is common and may have grave consequences in transplant recipients and congenitally infected children. Diagnosis of CMV infection is based on detection of specific antibodies and molecular assays. The incorporation of CMV serological assays into diagnostic...

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Published in:Journal of clinical microbiology Vol. 59; no. 8; p. e0096421
Main Authors: Miller, H, Simpson, P, Forman, M, Prigan, A, Kehl, S, Mesich, B, Faron, M, Ledeboer, N, Arav-Boger, R
Format: Journal Article
Language:English
Published: United States American Society for Microbiology 19-07-2021
Subjects:
Antibodies, Viral
Child
Cytomegalovirus - genetics
Cytomegalovirus Infections - diagnosis
Enzyme-Linked Immunosorbent Assay
Humans
Immunoassays
Immunoglobulin G
Membrane Glycoproteins
Viral Proteins
serotypes
UL144
cytomegalovirus
multiplex
ELISA
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Summary:Infection with human cytomegalovirus (CMV) is common and may have grave consequences in transplant recipients and congenitally infected children. Diagnosis of CMV infection is based on detection of specific antibodies and molecular assays. The incorporation of CMV serological assays into diagnostic algorithms requires careful evaluation and interpretation. Very few serological assays measure CMV infection by a specific strain. We developed an enzyme-linked immunosorbent assay (ELISA) using CMV-encoded UL144 as the antigen. UL144 encodes three major genotypes, A, B, and C, and recombinants. The ELISA was developed with the three UL144 proteins and optimized as a multiplex assay. Sera from 55 positive and 59 negative CMV IgG, determined by the clinical microbiology laboratory, were used for evaluation and optimization. A cutoff optical density (OD) that distinguishes UL144 antibody-positive from antibody-negative sera was established. UL144 A, B, C, and combinations of these antigens were detected in sera. An assay threshold of 0.1 was established and, from a total of 303 sera, the overall sensitivity, specificity, and positive and negative predictive values of the multiplex ELISA were 86.72% (95% confidence interval [CI] 79.59% to 92.07%), 96.57% (92.69% to 98.73%), 94.40% (88.45% to 97.38%), and 91.60% (87.50% to 94.44%), respectively. The inter- and intraassay median coefficients of variation were 0.06 (interquartile range [IQR] 0.56, 0.2) and 0.171 (IQR 0.038, 0.302), respectively. No cross-reactivity was observed with HSV-positive CMV-negative sera. This ELISA gives simple and reproducible results for detection of anti-CMV UL144 IgG. It may assist in differentiating natural infection from CMV vaccines that lack UL144, and may provide an important tool for epidemiological studies of CMV strains.
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Citation Miller H, Simpson P, Forman M, Prigan A, Kehl S, Mesich B, Faron M, Ledeboer N, Arav-Boger R. 2021. A novel multiplexed enzyme-linked immunosorbent assay for the detection of IgG seroreactivity to cytomegalovirus (CMV) UL144. J Clin Microbiol 59:e00964-21. https://doi.org/10.1128/JCM.00964-21.
ISSN:0095-1137
1098-660X
DOI:10.1128/JCM.00964-21