A novel flow cytometry-based tool for determining the efficiency of human cytomegalovirus infection in THP-1 derived macrophages

Huifen Li, Genxiang Mao, Joshua Carlson, Sean Leng

Research output: Contribution to journalArticle

Abstract

Human cytomegalovirus (hCMV) is a ubiquitous pathogen that causes congenital infection and severe infections in immunocompromised patients. Chronic hCMV infection may also play an important role in immunosenescence and adverse health outcomes in older adults. THP-1, a human monocytic cell line and its derived macrophages serve as a useful cell culture model for mechanistic studies of hCMV infection and its underlying biology. A major methodological challenge is the lack of a quick and reliable tool to accurately determine the efficiency of hCMV infection in THP-1 derived macrophages. In this study, we developed a flow cytometry based method using commercially available monoclonal antibody (MAb) against hCMV immediate early (IE) antigen that can accurately determine infection efficiency. We used 0.5% formaldehyde for fixation, 90% methanol for permeabilization, and incubation with FITC conjugated MAb at 37. °C. The method was tested by hCMV infection with laboratory Towne strain in the presence or absence of hydrocortisone. It was also compared with the routine flow cytometry protocol using Cytofix/Cytoperm solution and with immunofluorescence. The results indicate that this new method is reliable and time saving for accurate determination of infection efficiency. It may facilitate further investigations into the underlying biological mechanisms of hCMV infection.

Original languageEnglish (US)
Pages (from-to)127-130
Number of pages4
JournalJournal of Virological Methods
Volume221
DOIs
StatePublished - Sep 1 2015

Fingerprint

Cytomegalovirus Infections
Flow Cytometry
Macrophages
Infection
Monoclonal Antibodies
Fluorescein-5-isothiocyanate
Immunocompromised Host
Cytomegalovirus
Formaldehyde
Fluorescent Antibody Technique
Methanol
Hydrocortisone
Cell Culture Techniques
Cell Line
Health

Keywords

  • Flow cytometry
  • HCMV
  • Infection efficiency
  • THP-1

ASJC Scopus subject areas

  • Virology
  • Medicine(all)

Cite this

A novel flow cytometry-based tool for determining the efficiency of human cytomegalovirus infection in THP-1 derived macrophages. / Li, Huifen; Mao, Genxiang; Carlson, Joshua; Leng, Sean.

In: Journal of Virological Methods, Vol. 221, 01.09.2015, p. 127-130.

Research output: Contribution to journalArticle

@article{1cb1fad46726480da8f90ecd477da3ac,
title = "A novel flow cytometry-based tool for determining the efficiency of human cytomegalovirus infection in THP-1 derived macrophages",
abstract = "Human cytomegalovirus (hCMV) is a ubiquitous pathogen that causes congenital infection and severe infections in immunocompromised patients. Chronic hCMV infection may also play an important role in immunosenescence and adverse health outcomes in older adults. THP-1, a human monocytic cell line and its derived macrophages serve as a useful cell culture model for mechanistic studies of hCMV infection and its underlying biology. A major methodological challenge is the lack of a quick and reliable tool to accurately determine the efficiency of hCMV infection in THP-1 derived macrophages. In this study, we developed a flow cytometry based method using commercially available monoclonal antibody (MAb) against hCMV immediate early (IE) antigen that can accurately determine infection efficiency. We used 0.5{\%} formaldehyde for fixation, 90{\%} methanol for permeabilization, and incubation with FITC conjugated MAb at 37. °C. The method was tested by hCMV infection with laboratory Towne strain in the presence or absence of hydrocortisone. It was also compared with the routine flow cytometry protocol using Cytofix/Cytoperm solution and with immunofluorescence. The results indicate that this new method is reliable and time saving for accurate determination of infection efficiency. It may facilitate further investigations into the underlying biological mechanisms of hCMV infection.",
keywords = "Flow cytometry, HCMV, Infection efficiency, THP-1",
author = "Huifen Li and Genxiang Mao and Joshua Carlson and Sean Leng",
year = "2015",
month = "9",
day = "1",
doi = "10.1016/j.jviromet.2015.04.028",
language = "English (US)",
volume = "221",
pages = "127--130",
journal = "Journal of Virological Methods",
issn = "0166-0934",
publisher = "Elsevier",

}

TY - JOUR

T1 - A novel flow cytometry-based tool for determining the efficiency of human cytomegalovirus infection in THP-1 derived macrophages

AU - Li, Huifen

AU - Mao, Genxiang

AU - Carlson, Joshua

AU - Leng, Sean

PY - 2015/9/1

Y1 - 2015/9/1

N2 - Human cytomegalovirus (hCMV) is a ubiquitous pathogen that causes congenital infection and severe infections in immunocompromised patients. Chronic hCMV infection may also play an important role in immunosenescence and adverse health outcomes in older adults. THP-1, a human monocytic cell line and its derived macrophages serve as a useful cell culture model for mechanistic studies of hCMV infection and its underlying biology. A major methodological challenge is the lack of a quick and reliable tool to accurately determine the efficiency of hCMV infection in THP-1 derived macrophages. In this study, we developed a flow cytometry based method using commercially available monoclonal antibody (MAb) against hCMV immediate early (IE) antigen that can accurately determine infection efficiency. We used 0.5% formaldehyde for fixation, 90% methanol for permeabilization, and incubation with FITC conjugated MAb at 37. °C. The method was tested by hCMV infection with laboratory Towne strain in the presence or absence of hydrocortisone. It was also compared with the routine flow cytometry protocol using Cytofix/Cytoperm solution and with immunofluorescence. The results indicate that this new method is reliable and time saving for accurate determination of infection efficiency. It may facilitate further investigations into the underlying biological mechanisms of hCMV infection.

AB - Human cytomegalovirus (hCMV) is a ubiquitous pathogen that causes congenital infection and severe infections in immunocompromised patients. Chronic hCMV infection may also play an important role in immunosenescence and adverse health outcomes in older adults. THP-1, a human monocytic cell line and its derived macrophages serve as a useful cell culture model for mechanistic studies of hCMV infection and its underlying biology. A major methodological challenge is the lack of a quick and reliable tool to accurately determine the efficiency of hCMV infection in THP-1 derived macrophages. In this study, we developed a flow cytometry based method using commercially available monoclonal antibody (MAb) against hCMV immediate early (IE) antigen that can accurately determine infection efficiency. We used 0.5% formaldehyde for fixation, 90% methanol for permeabilization, and incubation with FITC conjugated MAb at 37. °C. The method was tested by hCMV infection with laboratory Towne strain in the presence or absence of hydrocortisone. It was also compared with the routine flow cytometry protocol using Cytofix/Cytoperm solution and with immunofluorescence. The results indicate that this new method is reliable and time saving for accurate determination of infection efficiency. It may facilitate further investigations into the underlying biological mechanisms of hCMV infection.

KW - Flow cytometry

KW - HCMV

KW - Infection efficiency

KW - THP-1

UR - http://www.scopus.com/inward/record.url?scp=84929617912&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84929617912&partnerID=8YFLogxK

U2 - 10.1016/j.jviromet.2015.04.028

DO - 10.1016/j.jviromet.2015.04.028

M3 - Article

VL - 221

SP - 127

EP - 130

JO - Journal of Virological Methods

JF - Journal of Virological Methods

SN - 0166-0934

ER -