摘要:
The present invention pertains to a method for efficiently introducing exogenous genes into primary lymphoid cells without drug selection which comprises the steps (a) deriving a retroviral vector and a helper cell combination that will yield a level of virus production in the range from 5.times.10.sup.6 to 5.times.10.sup.7 units/ml by transfecting a vector into a helper cell followed by selection, isolation of cell clones, and determination of viral titers to identify which virus-producing cell lines produce a virus titer in the range from 5.times.10.sup.6 to 5.times.10.sup.7 units/ml; (b) isolating a lymphoid cell subpopulation which can repopulate a specific lymphoid lineage or is a long-lived population by treating a suspension of lymphoid cells with a monoclonal antibody which removes undesired lymphoid cells to obtain an enriched lymphoid subpopulation; (c) culturing the enriched lymphoid subpopulation from step (b) with growth factors specific to the lymphoid subpopulation; (d) co-cultivating the lymphoid subpopulation from step (c) with a lawn of irradiated virus-producing cell line from step (a) to produce an infected lymphoid subpopulation; and (e) harvesting the infected lymphoid subpopulation.
摘要:
The present invention pertains to a method for efficiently introducing exogenous genes into primary lymphoid cells without drug selection which comprises the steps (a) deriving a retroviral vector and a helper cell combination that will yield a level of virus production in the range from 5.times.10.sup.6 to 5.times.10.sup.7 units/ml by transfecting a vector into a helper cell followed by selection, isolation of cell clones, and determination of viral titers to identify which virus-producing cell lines produce a virus titer in the range from 5.times.10.sup.6 to 5.times.10.sup.7 units/ml; (b) isolating a lymphoid cell subpopulation which can repopulate a specific lymphoid lineage or is a long-lived population by treating a suspension of lymphoid cells with a monoclonal antibody which removes undesired lymphoid cells to obtain an enriched lymphoid subpopulation; (c) culturing the enriched lymphoid subpopulation from step (b) with growth factors specific to the lymphoid subpopulation; (d) co-cultivating the lymphoid subpopulation from step (c) with a lawn of irradiated virus-producing cell line from step (a) to produce an infected lymphoid subpopulation; and (e) harvesting the infected lymphoid subpopulation. The invention further relates to a population of transfected lymphocytes, in which greater than about 90% of the lymphocytes contain a provirus.
摘要:
The present invention pertains to a method for efficiently introducing exogenous genes into primary lymphoid cells without drug selection which comprises the steps (a) deriving a retroviral vector and a helper cell combination that will yield a level of virus production in the range from 5.times.10.sup.6 to 5.times.10.sup.7 units/ml by transfecting a vector into a helper cell followed by selection, isolation of cell clones, and determination of viral titers to identify which virus-producing cell lines produce a virus titer in the range from 5.times.10.sup.6 to 5.times.10.sup.7 units/ml; (b) isolating a lymphoid cell subpopulation which can repopulate a specific lymphoid lineage or is a long-lived population by treating a suspension of lymphoid cells with a monoclonal antibody which removes undesired lymphoid cells to obtain an enriched lymphoid subpopulation; (c) culturing the enriched lymphoid subpopulation from step (b) with growth factors specific to the lymphoid subpopulation; (d) co-cultivating the lymphoid subpopulation from step (c) with a lawn of irradiated virus-producing cell line from step (a) to produce an infected lymphoid subpopulation; and (e) harvesting the infected lymphoid subpopulation. The invention further relates to a population of transfected lymphocytes, in which greater than about 90% of the lymphocytes contain a provirus.
摘要:
Isolated, latently infected T cell lines are provided that can be utilized in high throughput screening to discover compounds capable of activating HIV-I. The T cell lines harbor a latent HIV-I derived vector pro virus, which upon activation expresses a marker for late viral gene expression due to the insertion of the marker gene in the position of HIV-I envelope.
摘要:
Isolated, latently infected T cell lines are provided that can be utilized in high throughput screening to discover compounds capable of activating HIV-I. The T cell lines harbor a latent HIV-I derived vector pro virus, which upon activation expresses a marker for late viral gene expression due to the insertion of the marker gene in the position of HIV-I envelope.
摘要:
Novel packaging cell lines for production of lentiviral vectors are disclosed. The cell lines utilize an ecdysone-inducible system for virus production, thereby avoiding the cytotoxic effect of unregulated expression of certain envelope proteins and lentiviral proteins. Various embodiments of the cell lines are disclosed, which are suitable for different purposes that range from in vitro screening assays to gene therapy in vivo.
摘要:
Isolated, latently infected T cell lines are provided that can be utilized in high throughput screening to discover compounds capable of activating HIV-I. The T cell lines harbor a latent HIV-I derived vector pro virus, which upon activation expresses a marker for late viral gene expression due to the insertion of the marker gene in the position of HIV-I envelope.
摘要:
Isolated, latently infected T cell lines are provided that can be utilized in high throughput screening to discover compounds capable of activating HIV-I. The T cell lines harbor a latent HIV-I derived vector pro virus, which upon activation expresses a marker for late viral gene expression due to the insertion of the marker gene in the position of HIV-I envelope.
摘要:
A long-lived, myeloid-committed stem cell population is disclosed. Also disclosed are methods and compositions for targeting this population with retrovirus vectors in gene therapy protocols for correcting congenital disorders of myeloid system and for potentiating immune responses to defined tumor and viral antigens.