Optically Accessible Microfluidic Flow Channels for Noninvasive High-Resolution Biofilm Imaging Using Lattice Light Sheet Microscopy.
Optically Accessible Microfluidic Flow Channels for Noninvasive High-Resolution Biofilm Imaging Using Lattice Light Sheet Microscopy.
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DOI：
10.1021/acs.jpcb.1c07759
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发表时间：
2021-11-11
期刊：
影响因子：
--
通讯作者：
Gahlmann A
中科院分区：
文献类型：
--
作者：
Zhang J;Zhang M;Wang Y;Donarski E;Gahlmann A
需要能够对生物膜进行长期、高分辨率成像的成像平台来研究细菌生物膜内的细胞水平动力学。通过结合高空间和时间分辨率和低光毒性，晶格光片显微镜（LLSM）对细胞动力学的研究做出了重要贡献。然而，LLSM的力量尚未被用于生物膜研究，因为使用水浸物镜的开放式成像几何形状与活细菌标本不兼容;显微镜物镜上的细菌生长使得长期延时成像不可能，并为显微镜用户带来相当大的安全问题。为了使LLSM与致病性细菌标本兼容，我们开发了密封但光学可访问的微流体流动通道，可以在精确可控的物理和化学条件下维持细菌生物膜生长多日。为了产生液密和气密密封，我们在3D打印通道上粘了一层薄聚合物膜，其中顶壁被省略了。通过使用与水的折射率精确匹配的聚合物薄膜，我们实现了可忽略的光学像差。细菌本身不会粘附到聚合物膜上，因此聚合物窗口提供了通向通道内部的无阻碍的光学通路。在流动通道内，生物膜可以生长在任意的，甚至是不透明的表面上。通过将该流动通道与LLSM整合，我们能够以细胞分辨率记录几天内S. oneidensis MR-1生物膜的生长，而没有任何可观察到的光毒性或光损伤。
Imaging platforms that enable long-term, high-resolution imaging of biofilms are required to study cellular level dynamics within bacterial biofilms. By combining high spatial and temporal resolution and low phototoxicity, lattice light sheet microscopy (LLSM) has made critical contributions to the study of cellular dynamics. However, the power of LLSM has not yet been leveraged for biofilm research because the open-on-top imaging geometry using water-immersion objective lenses is not compatible with living bacterial specimens; bacterial growth on the microscope’s objective lenses makes long-term time-lapse imaging impossible and raises considerable safety concerns for microscope users. To make LLSM compatible with pathogenic bacterial specimens, we developed hermetically sealed, but optically accessible, microfluidic flow channels that can sustain bacterial biofilm growth for multiple days under precisely controllable physical and chemical conditions. To generate a liquid- and gas-tight seal, we glued a thin polymer film across a 3D-printed channel, where the top wall had been omitted. We achieved negligible optical aberrations by using polymer films that precisely match the refractive index of water. Bacteria do not adhere to the polymer film itself, so that the polymer window provides unobstructed optical access to the channel interior. Inside the flow channels, biofilms can be grown on arbitrary, even nontransparent, surfaces. By integrating this flow channel with LLSM, we were able to record the growth of S. oneidensis MR-1 biofilms over several days at cellular resolution without any observable phototoxicity or photodamage.
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DOI：
10.1126/science.aaq1392
发表时间：
2018-04-20
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Liu TL;Upadhyayula S;Milkie DE;Singh V;Wang K;Swinburne IA;Mosaliganti KR;Collins ZM;Hiscock TW;Shea J;Kohrman AQ;Medwig TN;Dambournet D;Forster R;Cunniff B;Ruan Y;Yashiro H;Scholpp S;Meyerowitz EM;Hockemeyer D;Drubin DG;Martin BL;Matus DQ;Koyama M;Megason SG;Kirchhausen T;Betzig E
通讯作者：
Betzig E
影响因子：
5.4
作者：
Grosskopf, Tobias;Soyer, Orkun S.
通讯作者：
Soyer, Orkun S.
DOI：
10.1007/s10577-021-09656-3
发表时间：
2021-03
期刊：
Chromosome research : an international journal on the molecular, supramolecular and evolutionary aspects of chromosome biology
影响因子：
--
作者：
Mimori-Kiyosue Y
通讯作者：
Mimori-Kiyosue Y
影响因子：
5.5
作者：
Manley, Harriet R.;Potter, David L.;Lieschke, Graham J.
通讯作者：
Lieschke, Graham J.
影响因子：
56.9
作者：
Gao, Ruixuan;Asano, Shoh M.;Betzig, Eric
通讯作者：
Betzig, Eric