Fluorescence Assays to Study Cell Adhesion and Migration In Vitro - Methods in Molecular Biology: Extracellular Matrix Protocols - page 323

20. For highly migratory cells incubation times can be rather short, i.e., 2-4 h,

whereas poorly migratory cells may require up to 72 h to accomplish significant
transmigrations. It is advisable to perform some pilot time-course studies to
empirically determine the optimal assay time for a given cell and/or experimental
condition. Kinetics of the transmigration/invasion process is possible when using
FATIMA plates ( Fig. 4 ), which allow for independent top and bottom fluores-
cence measurements. This time-related information can additionally be used as a
valuable parameter for establishing the migratory capacity of the cells.

21. The currently available membrane of the FATIMA plate shields the fluorescence

in the wavelength range of 450 to 550 nm. This implies that this plate can
optimally be employed when working with cells tagged with a fluorescent
label having emission spectra within that range, and hence, there is presently a
certain technical limitation when it comes to perform FATIMA experiments with
multiple labeled cells.

22. The FATIMA software is, similarly to the CAFCA one, a commercially available

(TECAN AG), Excel-based software that integrates the XFluor software needed
for the handling of the SPECTRAFluor Plus microplate fluorometer (TECAN
AG) and a specifically designed software for data elaboration.

Acknowledgments
The work was supported by TECAN Austria and grants from Consiglio
Nazionale delle Ricerche (CNR; p.f. ACRO), Associazione Italiana della
Ricerca sul Cancro (AIRC) and Fondo Sanitario Nazionale (Ricerca Finaliza-
tion 1995-1996). We are greatful to Johanna Neumayer and John Lipsky for
her numerous contributions to the development of the assays and Alfonso
Colombatti for his helpful suggestions.

References
1. DiMilla, P. A., Stone, J. A., Quinn, J. A., Albelda, S. M., and Lauffenburger, D. A.
(1993) Maximal migration of human smooth muscle cells on fibronectin and
type IV collagen occurs at an intermediate attachment strenght. J. Cell. Biol. 122,
729-737.
2. Palecek, S. P., Loftus, J. C., Ginsberg, M. H., Lauffenburger, D. G., and Horwitz, A. F.
(1997) Integrin-ligand binding properties govern cell migration speed through
cell-substratum adhesiveness. Nature 385, 537-540.
3. Roy, P., Petroll, W. M., Cavanagh, H. D., Chuong, C. J., and Jester, J. V. (1997)
An in vitro force measurement assay to study the early mechanical interaction
between corneal fibroblasts and collagen matrix. Exp. Cell Res. 232, 106-117.
4. Lotz, M. M., Burdsal, C. A., Erickson, H. P., and McClay, D. R. (1989) Cell
adhesion to fibronectin and tenascin: quantitative measurements of initial binding
and subsequent strengthening response. J. Cell. Biol. 109, 1795-1805.
5. Lallier, T. and Bronner-Fraser, M. (1991) Avian neural crest cell attachment to
laminin: Involvement of divalent cation dependent and independent integrins.
Development 113, 1069-1084.