Cells were imaged on a Zeiss Elyra 7 system using a Plan-Apochromat 40×/1.4 oil objective lens. Multicolored acquisition was performed sequentially to minimize cross-talk between channels. The fluorescent images were capture on two PCO Edge sCMOS cameras attached to a DuoLink motorized dual camera adapter and the color split using the secondary beam splitter BP420-480 + BP470-640 + LP740. The fluorescent protein mCherry was excited using the 561 nm laser line and emission collected from 570 to 640 nm. The fluorescent protein GFP was excited using the 488 nm laser line and emission collected from 490 to 570 nm. Apotome acquisition was set to collect five phase images with 25 ms camera exposure time. Yeast cells were optical Z sections using step size optimized for “Leap” acquisition. For time-lapse experiments z stacks were collected with an interval of 4.3 s. Apotome phase images were processed using Zeiss Zen Black software set to 3D SIM Leap. The alignment between the color channels was further improved by taking a Z stack of multicolor TetraSpec microspheres (ThermoFisher) that was used to generate an alignment matrix using the “Channel alignment” tool in Zen Black and applied to the time-lapse data.
Elyra 7 system
Manufactured by Zeiss
The Elyra 7 system is a high-performance imaging platform designed for advanced microscopy applications. It features a modular and configurable design, allowing users to tailor the system to their specific research needs. The Elyra 7 system enables high-resolution and high-speed imaging, providing researchers with powerful tools for studying complex biological samples.
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Lab products found in correlation
Tetraspec microspheres, by Thermo Fisher Scientific (2 mentions)
Plan apochromat 40 1.4 oil objective lens, by Zeiss (1 mentions)
Zen black, by Zeiss (1 mentions)
Arivis vision 4d, by Zeiss (1 mentions)
Alexa fluor 568, by Thermo Fisher Scientific (1 mentions)
Plan apochromat 63 1.4 oil objective, by Zeiss (1 mentions)
Prolong gold mounting media, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 647, by Thermo Fisher Scientific (1 mentions)
Zen black software, by Zeiss (1 mentions)
Plan apochromat 63x 1.4 oil objective, by Zeiss (1 mentions)
Fugene hd, by Promega (1 mentions)
6 protocols using elyra 7 system
1
Advanced Live-Cell Imaging with Zeiss Elyra 7
2
Super-resolution Imaging with Lattice SIM
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Super-resolution images were acquired with Elyra 7 system (lattice SIM2) from Zeiss, equipped with two cameras sCMOS. The objective used was 63× (NA 1.4) oil immersion and Z step was set between 0.094 and 0.110 μm. Image processing was based in SIM algorithm from ZEN 3.0 SR FP2 software from Zeiss. 3D processing was made using Arivis Vision4D also from Zeiss.
3
Structured Illumination Microscopy of Primary Neurons
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Primary neuronal cells were plated on high-precision type 1.5 coverslips. We incubated primary antibodies for 24 h at 4°C. We used Alexa secondary antibodies for each staining (IF: 1:500; Alexa Fluor 488, Alexa Fluor 568, Alexa Fluor 647; Molecular Probes, Thermo Fisher Scientific). These were incubated for 1 h 30 min at room temperature. We then mounted the coverslips on glass slides using ProLong Gold mounting media (Molecular Probes, Thermo Fisher Scientific). Our SIM acquisitions were done on a Zeiss Elyra 7 system equipped with lattice SIM technology. The signal was collected using a Zeiss Plan-Apochromat 63/1.4 Oil objective. We kept the laser power and exposure time to a minimum to avoid photobleaching. In the ZEN imaging software, the SIM mode was used to process the acquired images.
4
Super-Resolution Imaging of Bacterial Cell Walls
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Super-Resolution Structured Illumination Microscopy (SR-SIM) was performed to visualize cell wall staining in single bacterial cells. Briefly, bacteria were grown in THY at 37°C in 5% CO2 until midexponential phase. After washing with DPBS, bacteria were stained with CF 488-conjugated wheat germ agglutinin (WGA; Biotium) for 1 hour in the same incubation conditions. Bacterial cells were then washed and fixed with 4% (vol/vol) paraformaldehyde (PFA) solution for 20 minutes at 4°C. For the SR-SIM imaging, 10 µL of the sample was spotted on 1% agarose pads. The agarose pads were covered with No. 1.5H coverslips (Roth) and stored at 4°C for further imaging. The SR-SIM data were acquired on an Elyra 7 system (Zeiss) equipped with a 63×/1.4 NA Plan-Apochromat oil-immersion DIC M27 objective lens (Zeiss), a Piezo stage, and a PCO edge sCMOS camera with 82% QE and a liquid cooling system with 16-bit dynamic range. Using Lattice SIM mode, images were acquired with 13 phases. WGA CF 488 was detected with a 488-nm laser and a BP 495–590 emission filter. Super resolution images were computationally reconstructed from the raw data sets using default settings on ZenBlack software (Zeiss). Images were analyzed using the Fiji ImageJ software (77 (link)).
5
Simultaneous Imaging of Organelle Dynamics
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Coverglass, 8 Chambers from Eppendorf (0030742036) and incubated at 37°C and 5% CO2. After 24h cells were transfected with FuGENE® HD (Promega, E2311) in a ratio of 4:1 FuGENE (µl) : DNA (µg). Cells were incubated 24h -30h before imaging. Data of EMTB-3xGFP and tdTomato-Calreticulin was acquired simultaneously on a ZEISS Elyra 7 system with Duolink (two 4.2 CL HS pco.edge sCMOS cameras) using the 488 nm and 561 nm laser lines, Plan-Apochromat 63x 1.4 oil objective, 13 phases, 8 ms exposure time and 37°C. Simultaneous imaging allowed burst mode processing. mEmerald-Rab5a in U2OS cells was imaged with 9phases and 1 ms exposure time at 37°C.
6
Multicolor SIM Microscopy of Yeast Cells
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Cells were imaged on a Zeiss Elyra 7 system using Plan-Apochromat 40x/1.4 oil objective lens. Multi-coloured acquisition was performed sequentially to minimise cross talk between channels. The fluorescent images were capture on 2 PCO Edge sCMOS cameras attached to a DuoLink motorised dual camera adapter and the colour split using the secondary beam splitter BP420-480 + BP470-640 + LP740. The fluorescent protein mCherry was excited using 561nm laser line and emission collected from 570 to 640nm. The fluorescent protein GFP was excited using 488nm laser line and emission collected from 490 to 570nm. Apotome acquisition was set to collect 5 phase images with 25ms camera exposure time. Yeast cells were optical Z sections using step size optimised for "Leap" acquisition. For time lapse experiments z stacks were collected with an interval of 4.3 seconds. Apotome phase images were processed using Zeiss Zen Black software set to 3D SIM Leap. The alignment between the colour channels was further improved by taking a Z stack of multi-colour TetraSpec microspheres (ThermoFisher) that was used to generate an alignment matrix using the "Channel alignment" tool in Zen Black and applied to the time lapse data.
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