Calisesi Gianmaria, Castriotta Michele, Candeo Alessia, Pistocchi Anna, D'Andrea Cosimo, Valentini Gianluca, Farina Andrea, Bassi Andrea
Dipartimento di Fisica, Politecnico di Milano, piazza Leonardo da Vinci 32, 20133 Milano, Italy.
Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, via Fratelli Cervi 93, 20090 Segrate, Italy.
Biomed Opt Express. 2019 Oct 17;10(11):5776-5788. doi: 10.1364/BOE.10.005776. eCollection 2019 Nov 1.
Light sheet fluorescence microscopy has become one of the most widely used techniques for three-dimensional imaging due to its high speed and low phototoxicity. Further improvements in 3D microscopy require limiting the light exposure of the sample and increasing the volumetric acquisition rate. We hereby present an imaging technique that allows volumetric reconstruction of the fluorescent sample using spatial modulation on a selective illumination volume. We demonstrate that this can be implemented using an incoherent LED source, avoiding shadowing artifacts, typical of light sheet microscopy. Furthermore, we show that spatial modulation allows the use of Compressive Sensing, reducing the number of modulation patterns to be acquired. We present results on zebrafish embryos which prove that selective spatial modulation can be used to reconstruct relatively large volumes without any mechanical movement. The technique yields an accurate reconstruction of the sample anatomy even at significant compression ratios, achieving higher volumetric acquisition rate and reducing photodamage biological samples.
由于其高速度和低光毒性,光片荧光显微镜已成为最广泛使用的三维成像技术之一。三维显微镜的进一步改进需要限制样品的光照并提高体积采集率。我们在此提出一种成像技术,该技术允许使用选择性照明体积上的空间调制对荧光样品进行体积重建。我们证明这可以使用非相干LED光源来实现,避免了光片显微镜典型的阴影伪像。此外,我们表明空间调制允许使用压缩感知,减少要采集的调制图案数量。我们展示了斑马鱼胚胎的结果,证明选择性空间调制可用于在没有任何机械移动的情况下重建相对较大的体积。即使在显著的压缩率下,该技术也能准确重建样品解剖结构,实现更高的体积采集率并减少对生物样品的光损伤。