Scanning total internal reflection fluorescence imaging
Quirke AM., Ameer-Beg SM., Parsons M., Ng T., Irving M., Vojnovic B.
Cell adhesion and focal complex formation require signalling complexes linking cell adhesion molecules to the cytoskeleton. To understand morphogenetic changes associated with tumour cell spreading, migration and tumour cell metastasis, the molecular mechanisms responsible for the regulation, formation and dissolution at the cell-extracellular matrix (ECM) interface need to be identified. In order to achieve this, an improved axial resolution is desirable. We report on the development of a multi-photon (MP) total internal reflection (TIR) fluorescence lifetime imaging (FLIM) system that allows the selective excitation of fluorophores, with such an improved axial resolution. Results from initial experiments are presented. High excitation efficiency is achieved by the use of a Nikon 1.45 NA TIRF objective using annular illumination.