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Development of a microneutralization assay for HSV-2.
Horton, Melanie S; Minnier, Michael; Cosmi, Scott; Cox, Kara; Galli, Jennifer; Peters, Jessica; Sullivan, Nicole; Squadroni, Brian; Tang, Aimin; Fridman, Arthur; Wang, Dai; Chen, Zhifeng; Vora, Kalpit A.
Afiliación
  • Horton MS; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA. Electronic address: melanie.horton@merck.com.
  • Minnier M; AgileOne, Torrence, CA, USA.
  • Cosmi S; Eurofins Lancaster Laboratories Professional Scientific Service, Lancaster, PA, USA.
  • Cox K; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Galli J; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Peters J; Eurofins Lancaster Laboratories Professional Scientific Service, Lancaster, PA, USA.
  • Sullivan N; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Squadroni B; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Tang A; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Fridman A; Scientific Informatics, Merck & Co., Inc., Rahway, NJ, USA.
  • Wang D; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Chen Z; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
  • Vora KA; Infectious Diseases and Vaccines Discovery, Merck & Co., Inc., West Point, PA, USA.
J Virol Methods ; 297: 114268, 2021 11.
Article en En | MEDLINE | ID: mdl-34437874
BACKGROUND: Plaque Reduction Neutralization Test (PRNT) is the standard assay used for measuring neutralizing antibody responses to Herpes simplex virus type-2 (HSV-2). The PRNT is a cumbersome, time-consuming and laborious assay. The development of a faster, high throughput microneutralization assay (MNA) for HSV-2 viruses carried out in a 96-well format will allow for rapid testing of large numbers of samples for drug and vaccine development. METHODS: We describe the generation of a MNA that utilizes a pair of anti-HSV human monoclonal antibodies (mAbs) for virus detection in HSV-2 infected Vero cells. Antibodies were generated by B-cell cloning from PBMC's isolated from HSV-1 negative/HSV-2 positive donors. We describe the selection and characterization of the antibodies used for virus detection by ELISA with purified, recombinant anti-HSV glycoproteins, antibody binding in infected cells, and Western Blot. We determine the anti-HSV-2 neutralizing titers of immune sera from mice by MNA and PRNT and compare these results by linear regression analysis. RESULTS: We show that neutralization titers for HSV-2, determined by the 96-well MNA correlate with titers determined by a PRNT completed in 24-well plates in both the absence (R2 = 0.8250) and presence (R2 = 0.7075) of complement. CONCLUSIONS: We have successfully developed an MNA that can be used in place of the burdensome PRNT to determine anti-HSV-2 neutralizing activity in serum. This MNA has much greater throughput than the PRNT, allowing many more samples to be processed in a shorter time saving ∼90 % of the time required by the laboratory scientist to complete the task as compared to the traditional PRNT.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Herpesvirus Humano 2 / Anticuerpos Antivirales Límite: Animals Idioma: En Revista: J Virol Methods Año: 2021 Tipo del documento: Article Pais de publicación: Países Bajos
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
Buscar en Google

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Herpesvirus Humano 2 / Anticuerpos Antivirales Límite: Animals Idioma: En Revista: J Virol Methods Año: 2021 Tipo del documento: Article Pais de publicación: Países Bajos