Studying Cell–Surface Interactions In Vitro: A Survey of Experimental Approaches and Techniques - Tissue Engineering: Cell-Surface Interactions for Tissue Culture - page 47

Fig. 8 Detachment assays used to determine the mechanical resistivity in cell-substrate

interactions. a Centrifugation assay. b Hydrodynamic flow assay

more mechanical stress the cells can take without detachment, the more stable are
the cell-substrate interactions with the growth surface. Depending on the assay,
the mechanical stress is more or less defined and can be gradually increased to find
the critical shear force for detachment.
The simplest approach to study cell adhesion strength requires seeding of
suspended cells upon a substrate of interest. After a pre-defined incubation time,
the surface is rinsed with a physiological washing buffer. Weakly attached cells
adhering with a force smaller than the shear forces generated by the flow of buffer
are washed from the substrate surface. The number of cells that remain attached to
the surface is counted and serves as a measure for the adhesive interactions
between surface and cells under study. Although these wash-off assays provide
semi-quantitative information about the mechanical stability of cell-substrate
interactions, the applied shear forces are ill-defined, difficult to control and of
limited use [ 40 ].
Thus, more precise assays have been developed that use well-defined shear
forces to probe the stability of cell-surface interactions in mature cell populations.
According to the type of force application, these assays are classified as centri-
fugation assays [ 41 , 42 ] and hydrodynamic shear force assays [ 43 , 44 ].

4.2.1 Centrifugation Assay

The centrifugation assay (Fig. 8 a) applies the shear forces necessary to probe the
stability of cell adhesion by centrifugation—as the name implies. After cells are
allowed to fully attach and spread upon a surface under study, the cell-covered
surfaces are placed into centrifuge tubes filled with culture medium and are
centrifuged at a preset angular velocity. The centrifugal force acts as a well-
defined shear force parallel to the surface, generating tangential mechanical pull on
the cell bodies. After centrifugation the cells which resisted the mechanical stress
and remained attached to the substrate surface are counted using microscopic
techniques. Repeated runs with increasing angular velocity provide the critical
shear force that characterizes the mechanical stability of cell-substrate interactions