Adhesion and Structure

Adhesion plays an integral role in cell communication and regulation, and is of fundamental importance in the development and maintenance of tissues. Cell adhesion is involved in stimulating signals that regulate cell differentiation, the cell cycle, and cell survival. The adhesion of cells to each other or to the extracellular matrix (ECM) is responsible for a wide range of normal and aberrant cellular activity. This includes migration of immune cells to sites of infection, invasion and metastasis of tumor cells, and angiogenesis (the formation of new blood vessels) during wound healing. To perform many of these functions, cells must bind other cells or various molecules in the ECM.

Changes in cell adhesion can be the defining event in a wide range of diseases, including cancer, osteoporosis, atherosclerosis, and arthritis. The controlled regulation and the dynamics between integrins, MMPs, and ECM during tumor development and progression are also important areas of research.

Cell-extracellular matrix (ECM) interactions have global effects in many disease states through processes such as angiogenesis, apoptosis, and inflammation, and are critical for normal and tumor cell signal transduction pathways. The extracellular matrix is a complex structural and functional network of proteins and proteoglycans that can interact simultaneously with multiple cell surface receptors. A class of cell transmembrane proteins known as integrins mediate the adhesion of cells to ECM proteins and endothelial surfaces. These cell surface receptors anchor cells to the ECM leading to the transduction of signaling events that regulate cell survival, proliferation, and migration. Functional integrins are heterodimeric molecules, containing one a and one b subunit that are non-covalently bound together. Different integrin combinations may recognize a single ECM ligand, while others bind several different ECM proteins. Millipore provides a series of assays to facilitate discovery of the mechanisms and interactions that occur in the cellular adhesion process.

CytoMatrix™ Coated Strips

Convenient 12 x 8 stripwell format for flexibility in assay design
Consistent lot to lot coating concentration
Screening kit provides savings and profiling opportunities

Scientists are continually examining the adhesion and migration of many diverse cell types on various extracellular matrix (ECM) component proteins. CytoMatrix Cell Adhesion Strips are provided as 12 x 8-well removable strips in a plate frame for convenience and flexibility in designing assays. The wells in rows A–G have been coated with a human ECM protein. The wells in row H are coated with BSA to serve as a negative assay control. The CytoMatrix Screening Kit contains individual 96-well plates for fibronectin, vitronectin, laminin, collagen I and collagen IV. Cells are seeded onto the coated substrate. Subsequently, adherent cells are fixed and stained. Relative attachment is determined using absorbance readings.

Kit Components

ECM-coated Stripwell Plate
BSA-coated Control Well on each Stripwell

Integrin-Mediated & ECM Cell Adhesion Arrays

High throughput cell adhesion profiles in under two hours
Convenient 12 x 8 stripwell format for design flexibility
Colorimetric and fluorimetric formats available

Identification of cellular integrin profiles and ECM binding properties is the first step in understanding the mechanism of action in cell-ECM interactions. Historically, antibodies have been used to determine adhesion properties on a cell’s surface by immunoprecipitation, immunofluorescence, immunoblotting, or flow cytometry. These methods are laborious or require the use of sophisticated equipment. As an alternative, Millipore offers Integrin-Mediated Cell Adhesion Array kits and ECM Cell Adhesion Array kits as cost effective and efficient tools to screen cell surface protein profiles on virtually any human cell. Using the array format reduces variability in experimental conditions; therefore results are comparable in a consistent, controlled protocol. Kits are composed of stripwells with each well pre-coated with an individual integrin antibody or extracellular matrix protein and one negative control well coated with bovine serum albumin (BSA).

Kit Components

96-well Pre-coated Integrin or ECM Capture Plate
Built-in Negative Controls
Assay Buffer
Cell Stain Solution
Lysis Buffer (Fluorimetric kits)
Extraction Buffer (Colorimetric kits)

HL-60, HT-1080, KU812, RBL, and U937 cells were incubated for 2 hours at 37°C in wells coated with the array of a integrin monoclonal antibodies (Millipore cat. no. ECM530). The negative control well contains only the goat anti-mouse coating Ab. 100 µL of the cell suspension at 200,000 cells per well was added. After incubation, wells were washed, stained, and measured as described under Assay Instructions.

Capture Plate Stripwell Composition

Integrin-Mediated Cell Adhesion Kits

In addition to the integrin-mediated array formats, Millipore provides Integrin-Mediated Cell Adhesion kits with plates coated entirely of one integrin. These kits are useful to specifically screen the in vitro effects on the expression of one of these important integrins over an increased number of samples.

Cost effective with results in under two hours
Colorimetric assay does not require specialized instrumentation

Kit Components

96-well Pre-coated Integrin Capture Plate
Negative Control Plate
Cell Stain Solution
Extraction Buffer

3D Collagen Cell Culture Kit

Simulates collagen-rich ECM of normal tissues
Translucent collagen allows cells to be imaged within the matrix
Sterile and viable cells can be easily removed for further experimentation
All required reagents and media are included

The 3D Collagen Cell Culture Kit is a three dimensional cell culture system that closely simulates the collagen rich extracellular matrix of normal tissue. Cells suspended in this 3D system are easily visualized by phase contrast or fluorescence microscopy. Cells can be fixed and stained within the matrix and treated with antibodies for visualization of specific intra- and extracellular proteins. In addition, cells suspended in the 3D system can be treated with various reagents, allowing for extensive screening of biological responses to growth factors and chemical agents. Viable cells may be removed from the 3D cell culture system for further experimentation, including biological analysis and flow cytometry.

Kit Components

Collagen Solution RPMI Medium
M199 Medium
DMEM Medium
PBS with Phenol Red
Neutralization Solution

3D Collagen Cell Culture System Versus 2D Culture

	Confocal Image of Cos-7 actin cytoskeleton (red) in 3D Collagen Matrix. The nucleus is shown in purple.
	Actin cytoskeleton (red) of Cos-7 cells spread on 2D Collagen. The nucleus is show in purple.
	Apoptotic Cos-7 cells in 3D Collagen Matrix. The nucleus is shown in purple. Apoptotic bodies are shown in red.