Monitoring the dynamics of primary T cell activation and differentiation using long term live cell imaging in microwell arrays. Zaretsky, I., Polonsky, M., Shifrut, E., Reich-Zeliger, S., Antebi, Y., Aidelberg, G., Waysbort, N., & Friedman, N. Lab Chip, 12(23):5007–5015, Royal Society of Chemistry (RSC), December, 2012.
abstract   bibtex   
Methods that allow monitoring of individual cells over time, using live cell imaging, are essential for studying dynamical cellular processes in heterogeneous cell populations such as primary T lymphocytes. However, applying single cell time-lapse microscopy to study activation and differentiation of these cells was limited due to a number of reasons. First, primary naı̈ve T cells are non-adherent and become highly motile upon activation through their antigen receptor. Second, CD4(+) T cell differentiation is a relatively slow process which takes 3-4 days. As a result, long-term dynamic monitoring of individual cells during the course of activation and differentiation is challenging as cells rapidly escape out of the microscope field of view. Here we present and characterize a platform which enables capture and growth of primary T lymphocytes with minimal perturbation, allowing for long-term monitoring of cell activation and differentiation. We use standard cell culture plates combined with PDMS based arrays containing thousands of deep microwells in which primary CD4(+) T cells are trapped and activated by antigen coated microbeads. We demonstrate that this system allows for live cell imaging of individual T cells for up to 72 h, providing quantitative data on cell proliferation and death times. In addition, we continuously monitor dynamics of gene expression in those cells, of either intracellular proteins using cells from transgenic mice expressing fluorescent reporter proteins, or cell surface proteins using fluorescently labeled antibodies. Finally, we show how intercellular interactions between different cell types can be investigated using our device. This system provides a new platform in which dynamical processes and intercellular interactions within heterogeneous populations of primary T cells can be studied at the single cell level.
@ARTICLE{Zaretsky2012-hi,
  title     = "Monitoring the dynamics of primary {T} cell activation and
               differentiation using long term live cell imaging in microwell
               arrays",
  author    = "Zaretsky, Irina and Polonsky, Michal and Shifrut, Eric and
               Reich-Zeliger, Shlomit and Antebi, Yaron and Aidelberg, Guy and
               Waysbort, Nir and Friedman, Nir",
  abstract  = "Methods that allow monitoring of individual cells over time,
               using live cell imaging, are essential for studying dynamical
               cellular processes in heterogeneous cell populations such as
               primary T lymphocytes. However, applying single cell time-lapse
               microscopy to study activation and differentiation of these
               cells was limited due to a number of reasons. First, primary
               na{\"\i}ve T cells are non-adherent and become highly motile
               upon activation through their antigen receptor. Second, CD4(+) T
               cell differentiation is a relatively slow process which takes
               3-4 days. As a result, long-term dynamic monitoring of
               individual cells during the course of activation and
               differentiation is challenging as cells rapidly escape out of
               the microscope field of view. Here we present and characterize a
               platform which enables capture and growth of primary T
               lymphocytes with minimal perturbation, allowing for long-term
               monitoring of cell activation and differentiation. We use
               standard cell culture plates combined with PDMS based arrays
               containing thousands of deep microwells in which primary CD4(+)
               T cells are trapped and activated by antigen coated microbeads.
               We demonstrate that this system allows for live cell imaging of
               individual T cells for up to 72 h, providing quantitative data
               on cell proliferation and death times. In addition, we
               continuously monitor dynamics of gene expression in those cells,
               of either intracellular proteins using cells from transgenic
               mice expressing fluorescent reporter proteins, or cell surface
               proteins using fluorescently labeled antibodies. Finally, we
               show how intercellular interactions between different cell types
               can be investigated using our device. This system provides a new
               platform in which dynamical processes and intercellular
               interactions within heterogeneous populations of primary T cells
               can be studied at the single cell level.",
  journal   = "Lab Chip",
  publisher = "Royal Society of Chemistry (RSC)",
  volume    =  12,
  number    =  23,
  pages     = "5007--5015",
  month     =  dec,
  year      =  2012,
  language  = "en"
}
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