Comparison of dengue infection in human mononuclear leukocytes with mosquito C6/36 and mammalian Vero cells using flow cytometry to detect virus antigen

Sydow, Farid Fo von; Santiago, Marta A; Neves-Souza, Patrícia C.F.; Cerqueira, Denise; Gouvea, Adriana S; Lavatori, Maryrose Fh; Bertho, Álvaro Luiz; Kubelka, Claire Fernandes

doi:10.1590/s0074-02762000000400007

Cited by 24 publications

(17 citation statements)

References 17 publications

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“…It is unclear which cells are primary targets of dengue virus infection in vivo. In vivo studies suggest monocytes and lymphocytes are important targets, but these cell types are not highly susceptible to dengue virus infection in vitro (24,64,70). Dengue virus antigen has also been found in hepatocytes and endothelium from biopsies of dengue hemorrhagic fever patients (6,16,24,58).…”

Section: Resultsmentioning

confidence: 99%

Dengue Virus Nonstructural Protein NS5 Induces Interleukin-8 Transcription and Secretion

Medin¹,

Fitzgerald

Rothman

2005

J Virol

109

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Section: Resultsmentioning

confidence: 99%

Dengue Virus Nonstructural Protein NS5 Induces Interleukin-8 Transcription and Secretion

Medin¹,

Fitzgerald

Rothman

2005

J Virol

109

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“…Cells were stained with the anti-E-protein (pan-flavivirus) mouse mAb 4G2 conjugated to AlexaFluor-647 to measure intracellular expression of DENV E protein by flow cytometry. Detecting DENV antigen intracellularly by flow cytometry is a measure of viral replication (13). Inoculation of monocytes with DENV-2 alone caused little shift in DENV E-protein expression compared with mock conditions (Fig.…”

Section: Antibody-dependent Enhancement Of Dengue Virus Infection Incmentioning

confidence: 99%

“…When immune complexes are formed that do not meet the neutralization threshold, endocytosis of the complexes can promote increased cellular infection with DENV (9 -11). Circulating blood monocytes are thought to be the primary target of ADE in vivo and are largely resistant to DENV infection in the absence of enhancing antibodies (12)(13)(14)(15).…”

Section: Dengue Virus (Denv)mentioning

confidence: 99%

Spleen Tyrosine Kinase (Syk) Mediates IL-1β Induction by Primary Human Monocytes during Antibody-enhanced Dengue Virus Infection

Callaway

Smith

McKinnon

et al. 2015

Journal of Biological Chemistry

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Background: Insights into the mechanisms of elevated cytokine production during severe dengue disease are needed. Results: Dengue virus immune complexes induce inflammatory cytokine expression by activating spleen tyrosine kinase (Syk) during antibody-dependent enhancement of infection. Conclusion: Syk-mediated activation of ERK1/2 elevates cytokine production during antibody-enhanced dengue infection. Significance: Targeting Syk and ERK1/2 has therapeutic potential during antibody-enhanced dengue infection.

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“…Within the past 10 years, fluorescence-activated cell sorter (FACS)-based methods have been developed to follow infection and determine titers of viruses such as human immunodeficiency virus, herpesvirus, measles virus, influenza virus, Epstein-Barr virus, and rabies virus (1,4,12,13,16). FACS has also been used to detect DENV in clinical samples and to measure the ability of the virus to infect a variety of cells (2,3,7,8,11,17). Here we report on a FACS-based assay for titrating DENVs and for characterizing the ability of antisera to neutralize the virus.…”

mentioning

confidence: 99%

Flow Cytometry-Based Assay for Titrating Dengue Virus

et al. 2005

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Plaque assays for titrating dengue virus (DENV) are time-consuming and not suitable for strains that do not plaque. Fluorescence-activated cell sorting (FACS) has been used to detect DENV-infected cells. Here we describe a FACS-based assay for titrating DENV. We determined that at 24 h postinfection, the number of infected cells detected by FACS represented the first round of infection and therefore could be used as a readout of the number of infectious particles in the inoculum. When the titers of different laboratory and clinical strains of DENV were compared using FACS, plaque, and endpoint dilution assays, for most strains the FACS titers were comparable to titers obtained by plaque or endpoint dilution assays. The FACS assay is an improvement over the plaque assay because the infection period is reduced from 5 to 7 days to 24 h and the assay can be used to titrate clinical isolates that frequently do not form clear plaques on cell monolayers. The novel FACS-based methods described here will facilitate laboratory studies of dengue.Dengue is a mosquito-borne viral disease of global public health significance. Throughout the world there are more than 2.5 billion people at risk of dengue virus (DENV) infection. Each year there are an estimated 100 million cases of dengue viral infection worldwide, with 250,000 people developing the more severe dengue hemorrhagic fever/dengue shock syndrome, which is often fatal (5). DENV is a positive-polarity RNA virus in the family Flaviviridae, and the DENV complex consists of four distinct serotypes designated DENV1 through DENV4 (10).Laboratory studies of dengue virus are difficult because the virus does not grow to high titers in cell culture and assays for titrating virus and measuring virus neutralization are timeconsuming. These problems are exacerbated when working with primary clinical isolates of virus. Standard methods for titrating DENV and measuring the ability of antisera to neutralize the virus are based on plaque assays which require 5 to 7 days to complete. Some isolates, especially among primary clinical isolates, do not form clear plaques on cell monolayers. Better methods for titrating the virus and measuring the ability of antisera to neutralize dengue need to be developed. Within the past 10 years, fluorescence-activated cell sorter (FACS)-based methods have been developed to follow infection and determine titers of viruses such as human immunodeficiency virus, herpesvirus, measles virus, influenza virus, Epstein-Barr virus, and rabies virus (1,4,12,13,16). FACS has also been used to detect DENV in clinical samples and to measure the ability of the virus to infect a variety of cells (2,3,7,8,11,17). Here we report on a FACS-based assay for titrating DENVs and for characterizing the ability of antisera to neutralize the virus. MATERIALS AND METHODSCells, viruses, and antisera. Aedes albopictus C6/36 mosquito cells were obtained from the Centers for Disease Control and Prevention, Fort Collins. Vero 76 (African Green monkey kidney) cells were purchased fro...

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Comparison of dengue infection in human mononuclear leukocytes with mosquito C6/36 and mammalian Vero cells using flow cytometry to detect virus antigen

Abstract: Fluorescent activated cell sorter (FACS) analysis is useful for the detection of cellular

Cited by 24 publications

References 17 publications

Dengue Virus Nonstructural Protein NS5 Induces Interleukin-8 Transcription and Secretion

Dengue Virus Nonstructural Protein NS5 Induces Interleukin-8 Transcription and Secretion

Spleen Tyrosine Kinase (Syk) Mediates IL-1β Induction by Primary Human Monocytes during Antibody-enhanced Dengue Virus Infection

Flow Cytometry-Based Assay for Titrating Dengue Virus

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