-
超分辨结构光照明显微成像技术(super-resolution structured illumination microscopy,SR-SIM)具有时间分辨率高、光漂白和光毒性低和对荧光探针的要求少等优点，适用于活细胞的长时程超分辨成像。采用二维晶格结构光作为照明光，可以实现更快的成像速度和更低的光毒性，但同时也增加了系统的复杂性。为了解决此问题，本文提出了一种基于数字微镜器件的快速超分辨晶格结构光照明显微成像方法(lattice SIM based on a digital micromirror device,DMD-Lattice-SIM)，通过同步分时触发DMD和sCMOS相机的方式实现二维正交晶格结构光的产生，且只需要采集5幅相移原始图像即可重构出超分辨图像，相比于传统SR-SIM需要9幅相移原始图像的方法，图像采集时间减少了约44.4%。同时，在基于空域和频域联合的SIM重构算法(joint space and frequency reconstruction method-SIM,JSFR-SIM)的基础上，本文还发展了用于Lattice-SIM的JSFR超分辨图像重构方法(Lattice-JSFR-SIM)，先在频域对原始图像进行预滤波处理；然后，在空域对滤波后的图像进行超分辨重构处理。与传统频域图像重构处理对比，该方法，在512×512的成像视场下重构时间减少了约55.6%，对于实现活细胞实时超分辨成像具有重要意义和应用价值。Super-resolution structured illumination microscopy (SR-SIM) offers numerous advantages such high temporal resolution, low photobleaching and phototoxicity, and no special requirements for fluorescent probes. It is particularly suitable for long-term SR imaging of living cells. By using two-dimensional lattice structured light as illumination, SR-SIM can achieve faster imaging speed and reduced phototoxicity, albeit with increased system complexity. To address this challenge, this paper proposes a fast SR lattice structured illumination microscopy imaging method based on a digital micromirror device (DMD) called DMD-Lattice-SIM. This method utilizes a DMD and synchronous time-sharing triggering with sCMOS to generate two-dimensional orthogonal lattice structured light (As depicted in Figure 1). The proposed method only requires the collection of five phase-shifted raw images for SR image reconstruction, reducing the acquisition time by approximately 44.4% compared to the traditional SR-SIM method that requires nine phase-shifted raw images. Furthermore, this paper introduces a rapid SR image reconstruction method called Lattice-JSFR-SIM, which combines the benefits of joint space and frequency reconstruction (JSFR)-SIM and Lattice-SIM. The raw images are pre-filtered in the frequency domain and then undergo SR reconstruction in the spatial domain. This approach reduces reconstruction time by approximately 55.6% compared to traditional frequency domain image reconstruction processing, within an imaging field of view of 512×512. The feasibility of the proposed method is demonstrated through experiments on cell microtubules and the observation of mitochondrial division and fusion in living cells. The findings presented in this paper hold great significance and application value for enabling real-time SR imaging of living cells.
-
Keywords:
- structured illumination microscopy /
- super-resolution imaging /
- lattice structured illumination /
- spatial domain reconstruction
-
[1] Gustafsson M G L 2000 J. Microsc-oxford 198 82
[2] Chen X, Zhong S Y, Hou Y W, Cao R J, Wang W Y, Li D, Dai Q H, Kim D, Xi P 2023 Light-sci. Appl. 12 172
[3] Chang B J, Chou L J, Chang Y C, Chiang S Y 2009 Opt. Express 17 14710
[4] Dan D, Lei M, Yao B L, Wang W, Winterhalder M, Zumbusch A, Qi Y J, Xia L, Yan S H, Yang Y L, Gao P, Ye T, Zhao, W 2013 Sci. Rep. 3 1116
[5] Li M Q, Li Y N, Liu W H, Lal A, Jiang S, Jin D Y, Yang H P, Wang S, Zhanghao K, Xi P 2020 Appl. Phys. Lett. 116 233702
[6] Li X Y, Xie S Y, Liu W J, Jin L H, Xu Y K, Zhang L H, Hao X, Han Y B, Kuang C F, Liu X 2021 Opt. Express 29 43917
[7] Mudry E, Belkebir K, Girard J, Savatier J, Le Moal E, Nicoletti C, Allain M, Sentenac A 2012 Nat. Photonics 6 312
[8] Heintzmann R 2003 Micron 34 283
[9] Siebenmorgen J, Novikau Y, Wolleschensky R, Weisshart K, Kleppe I 2018 Introducing Lattice SIM for ZEISS Elyra 7 (Germany: Carl Zeiss Microscopy GmbH) pp1-7
[10] Zheng J J, Fang X, Wen K, Li J Y, Ma Y, Liu M, An S, Li J L, Zalevsky Z, Gao P 2022 Opt. Express 30 27951
[11] Huang X S, Fan J C, Li L J, Liu H S, Wu R L, Wu Y, Wei L S, Mao H, Lal A, Xi P, Tang L Q, Zhang Y F, Liu Y M, Tan S, Chen L Y 2018 Nat. Biotechnol. 36 451
[12] Zhao W S, Zhao S Q, Li L J, Huang X S, Xing S J, Zhang Y L, Qiu G H, Han Z Q, Shang Y X, Sun D E, Shan C Y, Wu R L, Gu L S, Zhang S W, Chen R W, Xiao J, Mo Y Q, Wang J Y, Ji W, Chen X, Ding B Q, Liu Y M, Mao H, Song B L, Tan J B, Liu J, Li H Y, Chen L Y 2022 Nat. Biotechnol. 40 606
[13] Wen G, Li S M, Wang L B, Chen X H, Sun Z L, Liang Y, Jin X, Xing Y F, Jiu Y M, Tang Y G, Li H 2021 Light-sci. Appl. 10 70
[14] Tu S J, Liu Q L, Liu X, Liu W J, Zhang Z M, Luo T J, Kuang C F, Liu X, Hao X 2020 Opt. Lett. 45 1567
[15] Dan D, Wang Z J, Zhou X, Lei M, Zhao T Y, Qian J, Yu X H, Yan S H, Min J W, Bianco P, Yao B L 2021 IEEE Photonics J. 13 3900411
[16] Wang Z J, Zhao T Y, Hao H W, Cai Y N, Feng K, Yun X, Liang Y S, Wang S W, Sun Y J, Bianco P R, Oh K, Lei M 2022 Adv. Photonics 4 026003
[17] Wang Z J, Zhao T Y, Cai Y A, Zhang J X, Hao H W, Liang Y S, Wang S W, Sun Y J, Chen T S, Bianco P R, Oh K, Lei M 2023 Innovation 4 100425
[18] Wen G, Li S M, Liang Y, Wang L B, Zhang J, Chen X H, Jin X, Chen C, Tang Y G, Li H 2023 PhotoniX 4 19
[19] Müller M, Mönkemöller V, Hennig S, Hübner W, Huser T 2016 Nat. Commun. 7 10980
[20] Zhou L L, Yu B, Huang L L, Cao H Q, Lin D Y, Jing Y Y, Wali F, Qu J L 2022 ACS Appl. Nano Mater. 5 18742
[21] Descloux A, Grussmayer K S, Radenovic A 2019 Nat. Methods 16 918
计量
- 文章访问数: 243
- PDF下载量: 11
- 被引次数: 0
下载:
