Scratch wound assay device

Details Scratch wound assay device Scratch wound assay device

2000-02-01

2001-10-30

Redding, David A. (Department: 1744)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

C435S309100, C600S570000, C600S306000

Reexamination Certificate

active

06309818

ABSTRACT:

TECHNICAL FIELD OF THE INVENTION
The invention relates generally to a tissue culture apparatus and methods of inducing a substantially reproducible wounding of cell, organ or tissue cultures for scratch wound assays.
BACKGROUND OF THE INVENTION
Conventional multiple well tissue culture plates as taught by Liner (U.S. Pat. No. 3,597,326) and Lyman (U.S. Pat. No. 4,012,288) are a common tool in most tissue culture experiments requiring small numbers of cells in relatively small volumes under multiple experimental conditions. The Terisaki tissue culture plate (U.S. Pat. No. 4,599,315) is used mainly in experiments aimed at cloning cells. Among the advantages of multiple well tissue culture plates are their low cost, disposable nature, and multiplicity of cell culture compartments. Multiple well tissue culture plates make possible multiple separate cultures all within a single culture device, thus making them attractive in screening assays. Multiple well tissue culture plates are available in standard configurations (e.g., 6, 12, 24, 48 or 96 wells per plate) which allow users to select a plate which has the desired well volume most suitable for a specific application. Plates with many wells, such as 48 or 96 well plates, are a good tool for use in experiments with many variables or experiments with a small number of cells. Plates with fewer numbers of wells that are larger in size are typically used in lieu of numerous separate cell culture flasks. It is the ability to handle a complete cell culture experiment with multiple parameters in an individual plate that makes these devices a common tool in most tissue culture experiments.
Previous studies have used a “scratch” wound closure assay to assess the potential effects of an agent on in vitro cell migration. However, presently available techniques result in non-reproducible model wounds which create difficulties in assessing the reproducibility of cellular migration and the dynamic process of wound repair.
SUMMARY OF THE INVENTION
The invention provides useful techniques, devices and systems for providing a substantially reproducible model wound in a tissue culture system to assess wound repair and cell migration.
The invention provides an apparatus having at least one template opening for causing a substantially reproducible injury to a cell, organ or tissue layer. The apparatus is useful in determining the effects of cell growth and migration agents in modeling wound closure produced in the cell, organ or tissue layer. The apparatus is useful in determining the effects of chemical, biochemical, biologic or bioengineered agents on cell growth and migration of any cell type. Also provided are methods of using the apparatus including juxtapositioning the apparatus to cells on a cell growth substrate and disrupting cells exposed through the template openings.
In one embodiment, a method of disrupting cells, comprising positioning in proximity a device including a template opening with a cell growth substrate having thereon a cell, organ or tissue layer; and exposing the cell, organ or tissue layer to a disruption medium through the template opening, wherein the disruption medium contacts the cells of the cell, organ or tissue layer; and disrupting the cells by contacting the cells with the disruption medium. The disruption medium can be any medium that is capable of causing injury to a cell, organ or tissue layer including photobleaching, laser ablation, UV exposure, thermal ablation, and mechanical disruption.
The cell culture growth substrate can be a multi-well tissue culture plate, a slide or planar surface. The template opening may be square, round, oblong or rectangular, but is typically longer than it is wide. In one embodiment the template opening is about 1.0 mm to about 2.0 mm wide and about 5 mm to about 9 mm in length.
In another embodiment, the invention provides an apparatus for use with a disruption medium for disrupting cells, the apparatus comprising a template opening to guide the disruption medium (e.g., a scratch device) and thereby limit disruption of the cells. The apparatus may contain at least one well, the well having at least one wall, a top and a bottom spaced apart from the top thereby defining a volume and having located therein the template opening.
In yet another embodiment, the invention provides an apparatus for guiding a scratch device during disruption of a cell, organ or tissue layer, the apparatus comprising a template opening sized to receive the scratch device and guide a tip thereof protruding from said template opening during disruption of a juxtaposed cell, organ or tissue layer.
In another embodiment, the invention provides a tray, comprising at least one well having an opening at the top and bottom and defining a first predetermined volume, and a template opening defining the bottom of the well, wherein a disruption medium may be directed through the template opening. The tray can be designed to be adaptably coupled to a reservoir, the reservoir defining a second predetermined volume greater than the first predetermined volume.


REFERENCES:
patent: 3597326 (1971-08-01), Liner
patent: 4012288 (1977-03-01), Lyman et al.
patent: 4599315 (1986-07-01), Terasaki et al.
patent: 5011779 (1991-04-01), Maimon

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