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Fluorescence-based quantitative methods for detecting human immunodeficiency virus type 1-induced syncytia
1Department of Internal Medicine, The University of Iowa, Iowa City, Iowa 52242, USA.
Insights
New fluorescence assays accurately detect and quantify human immunodeficiency virus type 1 (HIV-1) induced cell fusion (syncytia). These methods enable precise measurement of HIV replication and anti-HIV drug efficacy.
Area of Science:
- Virology
- Cell Biology
- Immunology
Background:
- Cell fusion, or syncytia formation, is a hallmark of human immunodeficiency virus type 1 (HIV-1) infection.
- Current methods for assessing syncytia lack quantification and do not estimate the number of cells involved in fusion.
Purpose of the Study:
- To develop and validate novel fluorescence-based methods for detecting and quantifying HIV-1-induced syncytia in vitro.
- To assess the utility of these assays for evaluating antiretroviral compounds.
Main Methods:
- Two fluorescence-based methods were employed: DNA staining with propidium iodide for flow cytometry analysis of cell size, and two-color cytoplasmic staining for fluorescence microscopy.
- Lymphoblastoid cell lines (MT-2, SupT1) were infected with syncytium-inducing (SI) HIV-1 isolates.
Main Results:
- Both fluorescence assays successfully detected and quantified HIV-induced syncytia.
- The methods differentiated between SI and non-SI HIV isolates and were applicable to multiple CD4(+) T-cell lines.
- Small syncytia were identifiable using the two-color cytoplasmic staining method.
- The assays accurately assessed the inhibitory effects of antiretroviral drugs like zidovudine and zalcitabine.
Conclusions:
- Fluorescence-based assays provide a rapid, practical, and quantitative approach to measure HIV-1 syncytia formation.
- These assays are valuable tools for studying HIV replication and screening potential anti-HIV compounds.
Abstract:
Cell fusion induced by human immunodeficiency virus type 1 (HIV-1) is usually assessed by counting multinucleated giant cells (syncytia) visualized by light microscopy. Currently used methods do not allow quantification of syncytia, nor do they estimate the number of cells involved in cell fusion. We describe two fluorescence-based methods for the detection and quantification of HIV-1-induced in vitro syncytium formation. The lymphoblastoid cell lines MT-2 and SupT1 were infected with syncytium-inducing (SI) HIV-1 isolates. Syncytia were detected by DNA staining with propidium iodide using flow cytometry to determine cell size or by two-color cytoplasmic staining of infected cell populations by using fluorescence microscopy. Both methods were able to detect and quantify HIV-induced syncytia. The methods could distinguish between SI and non-SI HIV isolates and could be used with at least two separate types of CD4(+) T-cell lines. Small syncytia can be readily identified by the two-color cytoplasmic staining method. Both methods were also shown to be useful for evaluating antiretroviral compounds, as demonstrated by the accurate assessment of HIV inhibition by azidothymidine (zidovudine), dideoxycytidine (zalcytibine), and hydroxyurea. These fluorescence-based assays allow a rapid and practical method for measuring HIV replication and anti-HIV activity of potential inhibitory compounds.
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