Schneider Jale, Aegerter Christof M
Physik-Institut, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland.
J Eur Opt Soc Rapid Publ. 2018;14(1):7. doi: 10.1186/s41476-018-0074-z. Epub 2018 Feb 21.
Light sheet microscopy became a popular tool allowing fast imaging with reduced out of focus light. However, when light penetrates turbid media such as biological tissues, multiple scattering scrambles the illumination into a speckle pattern and severely challenges conventional fluorescence imaging with focused light or with a light sheet. In this article, we present generation of light sheet type illumination patterns despite scattering.
We optimize the wave-front of the incoming light to transform the speckle pattern behind the scattering layer into a light sheet within the region of interest. We utilize a fast spatial light modulator for phase modulation and a genetic optimization algorithm. The light pattern behind the scattering layer is detected via a clear detection path and acts as a feedback signal for the algorithm.
We enabled homogenous light sheet illumination behind turbid media and enhanced the signal of fluorescent beads selectively at the desired focal plane up to eight times on average. The technique is capable to compensate the dynamic changes of the speckle pattern as well, as shown on samples consisting of living drosophila pupae.
Our technique shows that not only single foci, but also a homogenous light sheet illumination can directly be created and maintained behind static and dynamic scattering media. To make the technique suitable for common biological settings, where the detection path is turbid as well, a fluorescent probe can be used to provide the feedback signal.
光片显微镜成为一种流行的工具，可实现快速成像且减少离焦光。然而，当光穿透诸如生物组织等浑浊介质时，多次散射会将照明光散射成散斑图案，给传统的聚焦光或光片荧光成像带来严峻挑战。在本文中，我们展示了尽管存在散射仍能生成光片型照明图案。
我们优化入射光的波前，将散射层后的散斑图案在感兴趣区域内转换为光片。我们利用快速空间光调制器进行相位调制和遗传优化算法。通过清晰的检测路径检测散射层后的光图案，并将其用作算法的反馈信号。
我们实现了浑浊介质后方的均匀光片照明，并在所需焦平面上平均将荧光珠的信号增强了八倍。该技术还能够补偿散斑图案的动态变化，如在由活果蝇蛹组成的样本上所示。
我们的技术表明，不仅可以在静态和动态散射介质后方直接创建并维持单个焦点，还能实现均匀的光片照明。为使该技术适用于检测路径也浑浊的常见生物环境，可使用荧光探针来提供反馈信号。
