Multiphoton microscopy for blood vessel imaging: new non-invasive tools (Spectral, SHG, FLIM).

Imaging thick and opaque tissue, like blood vessel, in a noninvasive mode with high resolution, is nowadays possible with multiphoton technology. A near-infrared excitation presents the advantage to be compatible with living specimens and allows a deep penetration into tissues. The nonlinear excitation process is followed by several deactivation ways, among which fluorescence emission can be represented with Spectral or Lifetime imaging. Applied to ex vivo blood vessel imaging, these techniques enabled us to discriminate cell structures (nucleus, cytoskeleton) by fluorescent labelling (Hoechst, QDots). Another method, based on 2-photon excitation and which doesn't need any exogenous dye has also been experimented on arteries: SHG (Second Harmonic Generation) is a diffusion process generated from organized structures. Collagen molecules give rise to a strong SHG signal, enabling us to image the arterial wall (3-dimensional extracellular matrix).