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  Cell motility assayUSPTO Application #: 20060068374Title: Cell motility assay Abstract: An electrooptical method is disclosed for quantifying the motility response of cells to external stimuli, wherein a target cell population (formed as a function of said response) is subsequently optically differentiated by the addition of an optical differentiation solution, and whereby the targeted cell population becomes more or less detectable to an electrooptical reading device. The method can be used to perform chemotactic assays. For such and other purposes, the method can be performed utilizing high throughput robotic automation. (end of abstract) Agent: Nields & Lemack - Westboro, MA, US Inventor: Lakshmi Kamath USPTO Applicaton #: 20060068374 - Class: 435004000 (USPTO) Related Patent Categories: Chemistry: Molecular Biology And Microbiology, Measuring Or Testing Process Involving Enzymes Or Micro-organisms; Composition Or Test Strip Therefore; Processes Of Forming Such Composition Or Test Strip The Patent Description & Claims data below is from USPTO Patent Application 20060068374. Brief Patent Description - Full Patent Description - Patent Application Claims
[0001] This application is a divisional of U.S. Ser. No. 10/746,029 filed Dec. 23, 2003, the disclosure of which is incorporated herein by reference. FIELD [0002] In general, the present invention is directed to a cell motility assay, and in particular, a cell migration assay useful for studying cellular chemotaxis. BACKGROUND [0003] Cell movement, either as a chemotactic or chemokinetic response to soluble factors or as a phenomenon governed by a haptotactic response to cell surface or extracellular matrix-associated components, is one of the primary cellular processes during embryonic development, the maintenance of a healthy adult organism, and the progression of a number of pathological conditions, including cancer. [0004] Cell-based assays are widely implemented to study cell motility, and are particularly useful for pre-screening compounds to identify those compounds that can impair cell function and cell behavior. Cancer drug discovery efforts, for example, often incorporate chemotaxis-based cell motility assays to pre-screen and/or discover potential and promising drug candidates. [0005] Conventional chemotaxis assay procedures are well known. For example, according to one procedure, adherent cells are deposited onto one surface of an isoporous translucent polycarbonate membrane and placed within a solution containing a concentration gradient of pre-selected chemoattractant. Often, but not always, the chemoattractant is the subject of the investigation. The cells gradually move through the pores of membrane, from one surface to another, towards or away from higher or lower concentrations of the chemoattractant. The cells form migrant and non-migrant cell populations on respective sides of the membrane. The cells are then manually scraped off one surface of the membrane, and the remaining cells on the other surface are labeled with a fluorescent compound and "counted" with a scanning fluorometer. [0006] While this and like procedures are adequate for low throughput single-sample chemotactic assays (for example, for small-scale academic research), when seeking much higher throughput (for example, for large-scale industrial drug discovery efforts), the need to conduct several contemporaneous assays (for example, utilizing multi-segmented receptacles having several discrete assay chambers) becomes particularly acute. However, the difficulty of the physical steps involved in a single-sample assay are compounded when several instances of those steps are performed contemporaneously in (typically) smaller sample sizes. [0007] In particular, the step of removing non-migrant cells in a single chemotactic assay can be performed with comparative ease and efficacy. However, the same step conducted for ninety-six chemotactic samples, for example, plated within a 96-well multi-well array is difficult and prone to human error. More time, more effort, more care, and a higher level of technical skill is needed to manually scrape off cells. Often, each chamber must be individually and manually scrubbed, with each instance susceptible to cross contamination between the chambers and/or inadvertent membrane rupture. [0008] An alternative is suggested recently by U.S. Pat. No. 5,601,997, issued to R. Tchao on Feb. 11, 1997. [0009] The Tchao patent discloses a chemotactic methodology that utilizes a custom-engineered multi-well array as its assay sample platform. This multi-well array--unlike many comparable currently-available arrays--incorporates a "radiation opaque membrane". The radiation opaque membrane is disclosed as "having a plurality of substantially perpendicular pores" and is "not substantially transmissive" of "at least one" of certain defined wavelengths of electromagnetic radiation used in the course of assay. When an optical scanner is used for a fluorescence reading of labeled cells, the radiation opaque membrane blocks the passage of light, thus discouraging--as intended--both fluorescent activation and detection by the scanner of cells on the far opposite surface of the membrane. [0010] The Tchao patent provides an approach for conducting chemotactic assays that does not require manual cell removal prior to fluorescent scanning. Regardless, need remains for alternative approaches, particularly one that does not require the use of a custom-designed "radiation opaque membrane". SUMMARY [0011] Responsive to the above need, and other long felt needs, the present invention provides, in general, an electrooptical method for quantifying the motility response of cells to external stimuli, wherein a target cell population (formed as a function of said response) is subsequently optically differentiated by the addition of an optical differentiation solution, and whereby the targeted cell population becomes more or less detectable to an electrooptical reading device. [0012] The method can be conducted utilizing a comparatively broad variety of assay platforms (e.g., 96-well multi-well arrays), including "generic" transwell platforms. The method can also be conducted from commencement through completion, with neither any substantial physical disassembly of a utilized assay platform, nor any substantial intervening direct manual cell removal. The method is, for these and other factors, well-suited for high-throughput robotic automation. [0013] The method comprises particularly the steps of depositing cells (e.g., adherent tumor-causing cells) on a porous film (e.g., track-etched isoporous polycarbonate membrane); promoting migration of a portion of said cells from through said porous film (e.g., by establishing and placing the cells within a concentration gradient of a pre-selected chemoattractant); adding an optical differentiation solution to either the migrant or non-migrant cell portion to render said treated portion more or less detectable by an electrooptical reading device (e.g., a scanning fluorometer); then "counting" (directly or indirectly) either the migrant and/or non-migrant cell population utilizing said electrooptical reading device. [0014] In light of the above, it is a principal object of the present invention to provide an electrooptical method for quantifying the motility response of cells to external stimuli, wherein a target cell population (formed as a function of said response) is subsequently optically differentiated by the addition of an optical differentiation solution. [0015] It is another object of the present invention to provide a method for conducting a chemotactic assay that can be conducted utilizing a conventional electrooptical reading device and a generic chemotactic transwell assay platform (e.g., one without custom-made light blocking membranes). [0016] It is another object of the present invention to provide a method for conducting a chemotactic assay that can be conducted utilizing a conventional electrooptical reading device and a chemotactic transwell assay platform, and that is well-suited for robotic automation. [0017] It is another object of the present invention to provide a kit of pre-selected compatible components suitable for conducting pre-selected modes of practicing the inventive method. [0018] It is another object of the present invention to provide an optical attenuation solution useful for a comparatively broad range of modes of conducting the inventive method. [0019] These and other objects can be further appreciated in light of the detailed description herein of the invention according to certain of its current embodiments, considered together with the attached drawings. BRIEF DESCRIPTION OF THE DRAWINGS [0020] FIGS. 1a to 1d illustrate schematically a method for promoting, differentiating, and quantifying the chemotactic migration of cells C across a porous film 16, from one surface 16.sub.S to another 16.sub.D. Continue reading... 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