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TECHNIQUE TO MEASURE CELL MIGRATION

IP.com Disclosure Number: IPCOM000209218D
Publication Date: 2011-Aug-01
Document File: 5 page(s) / 45K

Publishing Venue

The IP.com Prior Art Database

Abstract

A fluorescence recovery after photobleaching (FRAP)-like technique to measure cell migration in two dimensions is disclosed. According to an embodiment of the invention, the number of cells migrating into a photobleached area provides a measure of cell migration or chemokinesis. The cell migration assay described herein is used in high throughput screening to analyze substances that regulate or inhibit cell migration.

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RP13422

TECHNIQUE TO MEASURE CELL MIGRATION

BRIEF ABSTRACT

    A fluorescence recovery after photobleaching (FRAP)-like technique to measure cell migration in two dimensions is disclosed. According to an embodiment of the invention, the number of cells migrating into a photobleached area provides a measure of cell migration or chemokinesis. The cell migration assay described herein is used in high throughput screening to analyze substances that regulate or inhibit cell migration.

KEYWORDS

    Fluorescence recovery after photobleaching, assay, high throughput screen, cell migration, chemokinesis.

DETAILED DESCRIPTION

    Conventional method of measuring cell migration involves a Boyden chamber. The Boyden chamber includes a culture insert containing 8 micrometer (┬Ám) pores where cells are placed. Cells that migrate through the pores adhere to the bottom of a membrane. Such cells adhering to the bottom of the membrane are measured or counted. Use of fluoroblok inserts modifies the Boyden chamber method for high throughput screens. The use of flouroblok inserts restricts fluorescent light from passing through the filter. The cells are fluorescently labeled and only the cells that migrate through the filter are measured with a fluorescence plate reader.

    Another conventional technique used to measure cell motility is phagokinetic tracks (PKT) assay. The PKT assay is performed on live cells plated on a lawn of microscopic fluorescent beads. The cells are labeled for 1


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RP13422

tracking purposes. Upon stimulation, the cells move across the lawn and phagocytose the beads, thereby, leaving phagokinetic tracks behind. The track area is proportional to the magnitude of cell movement.

    Yet another conventional cell migration assay provides a special device called a cell sedimentation manifold. The cell sedimentation manifold is developed for 1mm circular area of cells seeded on glass slides. The cells are seeded onto the top of the glass slides to form a circular area of cell culture. Random cell migration, or chemokinesis, makes the circular area bigger. The size of the circular area determines rate of cell migration. Further, steel barriers with an O ring are developed to seed cells in a defined circular area. Generally, the circular area has a diameter of ~7 millimeter (mm). The steel barriers are similar to the cell sedimentation chamber except that the steel barriers are used in multiwell plates (16mm wells).

    As illustrated in FIG. 1, Fluorescence Recovery After Photobleaching (FRAP) technology is , generally used to measure the mobility and/or diffusibilty of fluorescently labeled proteins in a cell. FRAP involves flashing an intense beam of light on a subsection of a cell containing fluorescently labeled molecules such that the fluorescent molecule is irreversibly modified and...