Microscopic and mesoscopic spectral bioimaging

Light microscopy has become a versatile tool for investigating biological phenomena as they unfold, using cells as living microcuvettes. The progress is based on improvements in a number of technological fields, including optics, electronics, reagent chemistry and computer science. The non-destructiveness, spatial resolution and speed of optical imaging can provide high versatility for investigating biological structure and function at the tissue or even whole organism level as well. Having previously reported on some new approaches to meeting the challenges posed by the fluorescence-based imaging of cells and tissues, we concentrate here on imaging with increased spectral content and resolution. We improved and applied two techniques of spectral selection: (1) acousto-optic tunable filtering, allowing for multiwavelength fluorescence microscopy with diffraction-limited spatial imaging and sub- millisecond temporal resolution, and (2) 2D Sagnac interferometry-based Fourier spectroscopy, yielding advanced spectral imaging capabilities. Enhanced implementations of existing optical technologies, coupled with image processing, were key in these approaches. We present an overview of the methods, and summarize some of our results in applying these advances to imaging biological specimens. The extension of spectral selection approaches to the mesoscopic domain, suitable for in vivo imaging is also illustrated, by fluorescence-based tumor visualization in the near infrared spectral region. Finally, some future directions are discussed.