Abstract
The kinetics of receptor-mediated cell adhesion to extracellular matrix and adherent cell monolayers plays a key role in many physiological and pathological processes including cancer metastasis. Within this process the presence of fluidic shear forces is a key regulator of binding equilibrium and kinetics of cell adhesion. Current techniques to examine the kinetics of cell adhesion are either performed in the absence of flow or are low throughput, limiting their application to pharmacological compound screening or the high throughput investigation of biological mechanisms. We developed a high throughput flow device that applies flow in a multi-well format and interfaced this system with electric cell-substrate impedance sensing (ECIS) system to allow label free detection of cell adhesion. We demonstrate that this combined system is capable of making real time measurements of cancer cell adhesion to extracellular matrix and immobilized platelets. In addition, we examined the dependence of the kinetics of binding of cancer cells on the level of shear stress and in the presence of small molecule inhibitors to adhesion-related pathways. This versatile system is broadly adaptable to the high throughput study of cell adhesion kinetics for many applications including drug screening and the investigation of the mechanisms of cancer metastasis.
Highlights
Parallel flow chamber assays can be performed with controlled steady flow and have found utility for measuring real time adhesion kinetics and rolling, but in most cases require observation with a microscope dramatically reducing throughput
The mechanisms of cancer cell adhesion during metastasis remain unclear and a major limitation in this area is a lack of high throughput assays for cell adhesion that can be carried out in the presence of the forces in circulatory flow
Our studies have validated the capability of a combined system that integrates a high throughput flow system and a label free detection system to perform accurate kinetic measurements of cancer cell adhesion under flow in the 96-well plate format
Summary
Parallel flow chamber assays can be performed with controlled steady flow and have found utility for measuring real time adhesion kinetics and rolling, but in most cases require observation with a microscope dramatically reducing throughput. Flow-incorporating assays would be useful for the development of small molecule inhibitors cell adhesion during cancer metastasis and a broad range of other physiological process. While many methods have been developed to precisely study cell adhesion kinetics, none are able to measure adhesion kinetics in the presence of shear stress in a high throughput manner. We have used this system to study the time course of adhesion of cancer cells to extracellular matrix and platelets. We performed high throughput studies of the effect of small molecule inhibitors on the adhesion kinetics of breast cancer cells
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