Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis
The human T-lymphotropic virus type-1 (HTLV-1) is an oncogenic retrovirus that predominantly spreads through cell-to-cell contact due to the limited infectivity of cell-free viruses. Among various modes of intercellular transmission, HTLV-1 biofilms emerge as adhesive structures, polarized at the ce...
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Bio-protocol LLC
2024-12-01
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| author | Coline Arone Hélène Dutartre Delphine Muriaux |
| author_facet | Coline Arone Hélène Dutartre Delphine Muriaux |
| author_sort | Coline Arone |
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| description | The human T-lymphotropic virus type-1 (HTLV-1) is an oncogenic retrovirus that predominantly spreads through cell-to-cell contact due to the limited infectivity of cell-free viruses. Among various modes of intercellular transmission, HTLV-1 biofilms emerge as adhesive structures, polarized at the cell surface, which encapsulate virions within a protective matrix. This biofilm is supposed to facilitate simultaneous virion delivery during infection. Yet, the molecular and functional intricacies of viral biofilms remain largely unexplored, despite their pivotal role in understanding retroviral pathogenesis. In this study, we optimized a protocol to isolate HTLV-1 biofilms from chronically infected T cells, facilitating their structural and molecular characterization using proteomic and super-resolution microscopy analyses. This protocol involves cultivating HTLV-1 chronically infected T cells at high density to facilitate the natural detachment of viral biofilms into the supernatant. Then, employing successive centrifugations, the cells are separated from the detached biofilms, and these structures are pelleted at medium speed (10,000× g). This method circumvents the need for mechanical, chemical, or enzymatic biofilm detachment, bypasses the use of ultracentrifugation, and enables us to resuspend the biofilms in the appropriate buffer for subsequent analyses such as western blotting or super-resolution microscopy imaging as presented. |
| format | Article |
| id | doaj-art-7f4a3c2494b44e38b8a0a0f08aab4187 |
| institution | Kabale University |
| issn | 2331-8325 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | Bio-protocol LLC |
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| series | Bio-Protocol |
| spelling | doaj-art-7f4a3c2494b44e38b8a0a0f08aab41872025-02-07T08:16:22ZengBio-protocol LLCBio-Protocol2331-83252024-12-01142410.21769/BioProtoc.5152Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity AnalysisColine Arone0Hélène Dutartre1Delphine Muriaux2Infectious Disease Research Institute of Montpellier (IRIM), UMR 9004 CNRS, University of Montpellier, Montpellier, FranceCenter for International Research on Infectiology (CIRI), UMR Inserm, ENS Lyon, Lyon, FranceInfectious Disease Research Institute of Montpellier (IRIM), UMR 9004 CNRS, University of Montpellier, Montpellier, FranceThe human T-lymphotropic virus type-1 (HTLV-1) is an oncogenic retrovirus that predominantly spreads through cell-to-cell contact due to the limited infectivity of cell-free viruses. Among various modes of intercellular transmission, HTLV-1 biofilms emerge as adhesive structures, polarized at the cell surface, which encapsulate virions within a protective matrix. This biofilm is supposed to facilitate simultaneous virion delivery during infection. Yet, the molecular and functional intricacies of viral biofilms remain largely unexplored, despite their pivotal role in understanding retroviral pathogenesis. In this study, we optimized a protocol to isolate HTLV-1 biofilms from chronically infected T cells, facilitating their structural and molecular characterization using proteomic and super-resolution microscopy analyses. This protocol involves cultivating HTLV-1 chronically infected T cells at high density to facilitate the natural detachment of viral biofilms into the supernatant. Then, employing successive centrifugations, the cells are separated from the detached biofilms, and these structures are pelleted at medium speed (10,000× g). This method circumvents the need for mechanical, chemical, or enzymatic biofilm detachment, bypasses the use of ultracentrifugation, and enables us to resuspend the biofilms in the appropriate buffer for subsequent analyses such as western blotting or super-resolution microscopy imaging as presented.https://bio-protocol.org/en/bpdetail?id=5152&type=0 |
| spellingShingle | Coline Arone Hélène Dutartre Delphine Muriaux Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis Bio-Protocol |
| title | Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis |
| title_full | Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis |
| title_fullStr | Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis |
| title_full_unstemmed | Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis |
| title_short | Isolation of Viral Biofilms From HTLV-1 Chronically Infected T Cells and Integrity Analysis |
| title_sort | isolation of viral biofilms from htlv 1 chronically infected t cells and integrity analysis |
| url | https://bio-protocol.org/en/bpdetail?id=5152&type=0 |
| work_keys_str_mv | AT colinearone isolationofviralbiofilmsfromhtlv1chronicallyinfectedtcellsandintegrityanalysis AT helenedutartre isolationofviralbiofilmsfromhtlv1chronicallyinfectedtcellsandintegrityanalysis AT delphinemuriaux isolationofviralbiofilmsfromhtlv1chronicallyinfectedtcellsandintegrityanalysis |