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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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.","journal":"Lab Chip","publisher":"Royal Society of Chemistry (RSC)","volume":"12","number":"23","pages":"5007–5015","month":"December","year":"2012","language":"en","bibtex":"@ARTICLE{Zaretsky2012-hi,\n title = \"Monitoring the dynamics of primary {T} cell activation and\n differentiation using long term live cell imaging in microwell\n arrays\",\n author = \"Zaretsky, Irina and Polonsky, Michal and Shifrut, Eric and\n Reich-Zeliger, Shlomit and Antebi, Yaron and Aidelberg, Guy and\n Waysbort, Nir and Friedman, Nir\",\n abstract = \"Methods that allow monitoring of individual cells over time,\n using live cell imaging, are essential for studying dynamical\n cellular processes in heterogeneous cell populations such as\n primary T lymphocytes. However, applying single cell time-lapse\n microscopy to study activation and differentiation of these\n cells was limited due to a number of reasons. First, primary\n na{\\\"\\i}ve T cells are non-adherent and become highly motile\n upon activation through their antigen receptor. Second, CD4(+) T\n cell differentiation is a relatively slow process which takes\n 3-4 days. As a result, long-term dynamic monitoring of\n individual cells during the course of activation and\n differentiation is challenging as cells rapidly escape out of\n the microscope field of view. Here we present and characterize a\n platform which enables capture and growth of primary T\n lymphocytes with minimal perturbation, allowing for long-term\n monitoring of cell activation and differentiation. 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