Light needles in scattering media using self-reconstructing beams and the STED principle

Abstract

Stimulated emission depletion (STED) microscopy generates super-resolved images of single cells by point-wise depletion of fluorescence around a small focal volume. Scanned light sheet microscopy, on the other side, generates images line-wise by scanning a weakly focused laser beam through thousands of scattering cells. Here we address the question of whether fluorescence from an excitation beam can be depleted by a STED beam over tens of micrometers while propagating through scattering material. Therefore, we use two self-reconstructing Bessel beams in continuous wave mode with a different angular momentum for fluorescence excitation and depletion along a distance of 110 μm. We show that despite significant scattering at various arrangements of microspheres embedded in agarose gel and despite strong losses in spatial coherence, it is possible to generate a sufficiently good overlap of both beam intensities. Without affecting the self-healing capability of the illumination photons in the Bessel beam’s ring system, the emission of fluorescence photons thereof can be strongly suppressed. This results in a needle-like fluorescence distribution inside scattering media, providing new perspectives for fundamental principles and applications in microscopy and metrology.