Chemotaxis materials

We generally assess the protein concentration of new chemokine stock solutions by OD280 using the appropriate extinction coefficient for each chemokine or by Bradford assay. Although this is not often practical when very small quantities of chemokine are purchased because 10-20% of the material may be sacrificed for an accurate determination, we have found that the actual protein concentration may vary by as much as 2CM-0% between lots which can significantly effect the reproducibility of chemotaxis assays. 

The Boyden chamber is a simple apparatus used to test for chemotaxis, especially of leukocytes. It can also be used to assess tumor cell transmigration across an endo-thehal monolayer in vitro. It consists of two compartments separated by a MiUi-pore filter (3-8 pm pore size). A chemotac-tic factor is placed in one compartment, and a gradient develops across the thickness of the filter (ca. 150 pm). Cell movement into the filter is measured after an incubation period less than the time taken for the gradient to decay. Cell motility can be measured in Boyden chambers containing filters precoated with different materials, for example fibronectin or fibronectin fragments. The method, when apphed to malignant and non-mahgnant cell hnes, shows that the variable invasive potentials of these cells correlate with their abihty to disrupt the endothelial cell monolayer. 

The following sections integrate the material described above to provide our current understanding of the mechanisms involved in Dictyostelium and mesenchymal cell chemotaxis. Dictyostelium vwll be covered first, and provides specific paradigms for spatial discrimination that are relatively well worked out. How these paradigms could apply to mesenchymal cell chemotaxis is then discussed in the subsequent section along with additional pathways. 

Chemotaxis Assay for Differentiated HL-60 Ceiis with Gradient Switch Materials ... 

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