Extended focus Fourier domain optical coherence microscopy and fluorescence lifetime imaging

Fourier domain Optical coherence microscopy (FDOCM) offers excellent sensitivity and high axial resolution to image the structure of biological tissue. The depth information is extracted in parallel and allows very high volume acquisition rates. An illumination scheme employing an axicon lens provides extended depth of field (xf) in combination with high lateral resolution. The xf-OCM signal reveals the spatial distribution of changes of the refractive index in the sample with high contrast at a near isotropic resolution of ~2um. However identification and differentiation between different tissues may be difficult with this purely structural sample information at hand. Combining the xf-OCM setup with fluorescence lifetime imaging (FLIM) offers a powerful method to obtain functional contrast with molecular specificity. The FLIM- system works in the Fourier domain measuring the phase offset between the modulated excitation signal and the returned fluorescence intensity. Both the fluorescence amplitude and lifetime are retrieved. Previously labeled structures can be identified and help interpreting the xf- OCM signal. The fluorescence lifetime provides additional functional information. It varies between different fluorophores and can be very sensitive to environmental changes. The xf-OCM tomograms and FLIM images are acquired in parallel and provide a complementary view of the sample. This combined system was applied to the imaging of biological tissue such as rat hair follicle and mouse pancreas.