Cellomics HCS Reagent Kits for multiplex analysis of cytoskeletal parameters: F-actin (microfilament) and tubulin (microtubule) structure, plus nuclear and whole cell morphology.

Description

The Cytoskeletal Rearrangement HCS Reagent Kits are for the simultaneous quantitation of DNA content, cell morphology, and the intracellular arrangement of microfilaments and microtubules in the same cell (Figure 1). These kits allow direct measurements of cell and nuclear morphology, F-actin, and microtubule changes using a fixed end-point assay based on immunofluorescence detection in cells grown on standard high-density microplates. The primary antibody is specific for its target and has minimal cross-reactivity with other targets.

These kits have been optimized with the Cellomics ArrayScan HCS Reader using the Morphology BioApplication Software Module. Thus, automated plate-handling, focusing, cell image acquisition/processing, and data analysis/management are combined in one high-content screening (HCS) system to assay for test compounds. In addition to HCS instruments, cells labeled by the kit reagents can be viewed and analyzed by other fluorescence microscopes.

NIH3T3 cells	HMVEC-L cells
Figure 1. Staining of cytoskeleton structure in NIH3T3 cells (left, 20X objective) and HMVEC-L cells (right, 20X objective). Markers were detected according to the kit protocol. Cell images were acquired using the Cellomcs ArrayScan HCS Reader. Colors are F-actin (red), tubulin (green), nuclei stained with DAPI (blue).

Background

The intracellular meshwork of the cytoskeleton is responsible for maintaining cell shape, cell movement, cytokinesis and organelle organization.^1,2 The cytoskeleton network also facilitates proper function of other proteins by direct binding, transporting, repositioning and sequestering these proteins. The structure of cytoskeleton is controlled by cytoskeleton-associated proteins in response to the external signaling.³ Therefore, defects in the ability to regulate the dynamics of cytoskeletal structure are likely to cause detrimental effects on other cell function.⁴

Cytoskeletal rearrangement is often associated with cellular toxicity, pathology and cell death.^4,5 Signaling defects in conjunction with cytoskeletal rearrangement can also contribute to cell proliferation and tumor cell activation, which result in metastasis.^6,7 An accurate assay for cytoskeletal rearrangement is essential to evaluate potential therapeutic targets.

High-content analysis (HCA) involves a fluorescence cell-based assay in which cells are automatically imaged and analyzed using fluorescence microscopy. The Multiplexed Cytoskeletal Rearrangement HCS Reagent Kits provide a highly effective tool for studying the cytoskeletal changes. The reagents combined with the Cellomics ArrayScan HCS Reader and the Morphology BioApplication Software Module allows accurate quantification of microfilaments and microtubules simultaneously within the same cell (Figure 2).

See product instructions for additional example data and literature references.

Non-treated	Cytochalasin D	Staurosporine
Figure 2. Staining of DAPI (nucleus, blue), F-actin (microfilament, red) and tubulin (microtubule, green) in NIH 3T3 cells. Cells were treated with 10 μM cytochalasin D or 1 μM staurosporine for 3 hours or incubated only with media (non-treated). Markers were detected according to the kit protocol. Cytochalasin D and staurosporine dismantle cytoskeleton structure, which causes dramatic morphological change in cells. The cell images were acquired using a Cellomics ArrayScan HCS Reader. Graphs (thumbnail images in bottom row) are dose response curves for NIH3T3 cell data representing mean +/-SD from three plates (sixteen wells per point in a plate) and analyzed using the Morphology BioApplication Software Module.

Related Poster Presentations

	Suk J. Hong and Richik N. Ghosh (2008) Quantitative Analysis of Drug Effects on Cytoskeletal Structure, Cell Morphology and the Cell Cycle by High-Content Assays.
	Suk J. Hong, Richik N. Ghosh (2007) Simultaneous, Quantitative Monitoring of Cytoskeleton Rearrangement and Morphology Changes in Cells Using High Content Analysis.
	Suk J. Hong, Michael Anhalt, and Richik N.Ghosh (2007) New High Content Screening Reagent Kits Enabling Simultaneous Quantitative Monitoring of Cytoskeleton Rearrangement and Cell Morphology Changes.
	B. Nickel, C. Paroz, C. Falk, P Zbinden, U. Regenass (2006) High Content Screening for Apoptosis in Drug Discovery.
	Richik N. Ghosh, Michael A. Sipe, Linnette E. Grove, Rich Jarvis, Mike Byrom, David Brown & Jeffrey R. Haskins (2005) Identification of Kinases Affecting Downstream Phenotypic Changes in Cell Morphology and Cytoskeleton by the High Content Screening Evaluation of a siRNA Library.
	Linnette Grove, Richik N. Ghosh, Oleg Lapets and Everett Ramer (2004) Automated Quantitative Analysis of Cytoskeletal Rearrangement Over Multiple Spatial Dimensions Using High Content Screening.
	Sarah Burroughs Tencza and Jeffrey R. Haskins (2004) High-Content Profiling of Cell Cycle Related Targets with RNAi.
	Richard DeBiasio, Richik N. Ghosh, Alexander L. Friedman and Jeffrey R. Haskins (2004) Optical Sectioning With the Apotome Attachment on the ArrayScan HCS Reader Enhances High Content Screening Assays.
	Cellomics, Inc. (2003) Unlocking the Power of Optical Sectioning with the ArrayScan V HCS Reader.

Kit Contents

Ordering Information

Screening size kits and components (bulk and custom) available with special pricing. Please call.

To order Cellomics HCS Reagent Kits, please call 800-874-3723 in the U.S.
or contact your local distributor of Thermo Scientific Products.

Choose "Order Online" in table to see pricing and order online (U.S. only).

Other Whole Cell Stains are also available: see Cellomics HCS Reagent Kits.