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Cell Microharpooning to Study Nucleo-Cytoskeletal Coupling.

ABSTRACT: To evaluate the intracellular force transmission between the nucleus and cytoskeleton, we optimized a single cell-based assay that involves the manipulation of living, adherent cells with a fine glass microneedle and a microscope-mounted micromanipulator. The user inserts the microneedle into the cytoplasm and then, using a custom-programmable computer script, pulls the needle laterally toward the cell periphery. Normalized cross-correlation is applied to recorded time-lapse image sequences to determine average displacements within predefined regions of the nucleus and the cytoskeleton. These regional displacements, together with calculations of nuclear elongation, nuclear centroid translocation, and nuclear shape changes, enable quantitative assessments of nucleo-cytoskeletal coupling in both normal and disease conditions and provide an improved understanding of the role of specific nuclear envelope proteins in intracellular force propagation.

SUBMITTER: Fedorchak G

PROVIDER: S-EPMC5966280 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Cell Microharpooning to Study Nucleo-Cytoskeletal Coupling.

Fedorchak Gregory G Lammerding Jan J

Methods in molecular biology (Clifton, N.J.) 20160101

To evaluate the intracellular force transmission between the nucleus and cytoskeleton, we optimized a single cell-based assay that involves the manipulation of living, adherent cells with a fine glass microneedle and a microscope-mounted micromanipulator. The user inserts the microneedle into the cytoplasm and then, using a custom-programmable computer script, pulls the needle laterally toward the cell periphery. Normalized cross-correlation is applied to recorded time-lapse image sequences to d ...[more]

PMID: 27147047

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