Development of a high-throughput β-Gal-based neutralization assay for quantitation of herpes simplex virus-neutralizing antibodies in human samples.
Measurement of neutralizing antibodies against herpes simplex virus (HSV) is important for evaluation of candidate vaccines. The established plaque-reduction neutralization assay is time consuming, labor intensive, and difficult to validate and transfer. Here, we describe the characterization of a HSV-neutralization assay based on the expression of a reporter gene, β-galactosidase (β-Gal). Using previously constructed HSV-β-Gal recombinant viruses, HSV-2/Gal and HSV-1/tk12, we developed a colorimetric β-Gal-based neutralization assay that is sensitive and highly reproducible, and performed in less than 48h. HSV-1 and HSV-2 neutralizing titers measured by the β-Gal-based neutralization assay were equivalent to those obtained by a plaque reduction neutralization assay. Intra- and inter-assay precision studies demonstrated that the β-Gal-based assay was repeatable and yielded low and acceptable variation. In addition, comparison of HSV-2 neutralizing antibody (NAb) titers measured in two independent laboratories by two unique β-Gal-based assays showed a highly significant correlation (r=0.9499, p<0.0001) between the two assays. The new assay will serve as an important tool both for preclinical and clinical trials of new HSV vaccines.
Genocea Biosciences, Cambridge, MA 02140, United States.
Amy
A
Baccari
Genocea Biosciences, Cambridge, MA 02140, United States.
Michael
M
Cooney
Dept. of Internal Medicine, Saint Louis University School of Medicine, St. Louis, MO 63104, United States.
Tamara P
TP
Blevins
Dept. of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, MO 63104, United States.
Lynda A
LA
Morrison
Genocea Biosciences, Cambridge, MA 02140, United States.
Shane
S
Larson
Genocea Biosciences, Cambridge, MA 02140, United States.
Mojca
M
Skoberne
Dept. of Internal Medicine, Saint Louis University School of Medicine, St. Louis, MO 63104, United States.
Robert B
RB
Belshe
Genocea Biosciences, Cambridge, MA 02140, United States. Electronic address: jessica.flechtner@genocea.com.
Jessica B
JB
Flechtner
Genocea Biosciences, Cambridge, MA 02140, United States.
Deborah
D
Long
D000818:Animals; D057134:Antibodies, Neutralizing / Q000097:blood*; D000914:Antibodies, Viral / Q000097:blood*; D002522:Chlorocebus aethiops; D017930:Genes, Reporter; D006561:Herpes Simplex / Q000097:blood / Q000276:immunology*; D018259:Herpesvirus 1, Human; D018258:Herpesvirus 2, Human; D057166:High-Throughput Screening Assays; D006801:Humans; D009500:Neutralization Tests; D015203:Reproducibility of Results; D014709:Vero Cells; D001616:beta-Galactosidase / Q000235:genetics
D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D001616:beta-Galactosidase
Antibody titer; Assay; Herpes simplex virus; Neutralization