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In general, it is assumed that the reservoir consists of CD4\n            <jats:sup>+<\/jats:sup>\n            T cells that express no viral proteins. However, recent findings suggest that this may be an overly simplistic view and that the cells that contribute to the reservoir may be a diverse population that includes both CD4\n            <jats:sup>+<\/jats:sup>\n            and CD4\n            <jats:sup>\u2212<\/jats:sup>\n            cells. In this study, we directly infected resting CD4\n            <jats:sup>+<\/jats:sup>\n            T cells and used fluorescence-activated cell sorting (FACS) and fiber-optic array scanning technology (FAST) to identify and image cells expressing HIV Gag. We found that Gag expression from integrated proviruses occurred in resting cells that lacked surface CD4, likely resulting from Nef- and Env-mediated receptor internalization. We also extended our approach to detect cells expressing HIV proteins in patients suppressed on ART. We found evidence that rare Gag\n            <jats:sup>+<\/jats:sup>\n            cells persist during ART and that these cells are often negative for CD4. We propose that these double-negative \u03b1\/\u03b2 T cells that express HIV protein may be a component of the long-lived reservoir.\n          <\/jats:p>\n          <jats:p>\n            <jats:bold>IMPORTANCE<\/jats:bold>\n            A reservoir of infected cells persists in HIV-infected patients during antiretroviral therapy (ART) that leads to rebound of virus if treatment is stopped. In this study, we used flow cytometry and cell imaging to characterize protein expression in HIV-infected resting cells. HIV Gag protein can be directly detected in infected resting cells and occurs with simultaneous loss of CD4, consistent with the expression of additional viral proteins, such as Env and Nef. Gag\n            <jats:sup>+<\/jats:sup>\n            CD4\n            <jats:sup>\u2212<\/jats:sup>\n            cells can also be detected in suppressed patients, suggesting that a subset of infected cells express proteins during ART. Understanding the regulation of viral protein expression during ART will be key to designing effective strategies to eradicate HIV reservoirs.\n          <\/jats:p>","DOI":"10.1128\/jvi.01913-15","type":"journal-article","created":{"date-parts":[[2015,11,5]],"date-time":"2015-11-05T03:43:32Z","timestamp":1446695012000},"page":"2165-2179","update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":65,"title":["A Subset of CD4\/CD8 Double-Negative T Cells Expresses HIV Proteins in Patients on Antiretroviral Therapy"],"prefix":"10.1128","volume":"90","author":[{"given":"Laura K.","family":"DeMaster","sequence":"first","affiliation":[{"name":"Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaohe","family":"Liu","sequence":"additional","affiliation":[{"name":"Center for Cancer and Metabolism, SRI International, Menlo Park, California, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"D. Jake","family":"VanBelzen","sequence":"additional","affiliation":[{"name":"Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benjamin","family":"Trinit\u00e9","sequence":"additional","affiliation":[{"name":"Department of Basic Science, New York University College of Dentistry, New York, New York, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lingjie","family":"Zheng","sequence":"additional","affiliation":[{"name":"Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Luis M.","family":"Agosto","sequence":"additional","affiliation":[{"name":"Section of Infectious Diseases, Boston University School of Medicine and Boston Medical Center, Boston, Massachusetts, 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