For semi-quantitative analyses of 6-OHDA-induced degeneration, worms were examined using a Leica fluorescent dissecting microscope. The absence of all eight dopaminergic neurons in worms was scored as “complete loss.” The presence of a complete, intact set of eight dopaminergic neurons was scored as “no loss.” Any intermediate situation, for example a damaged or absent subset of dopaminergic neurons or missing dendrite portions, was scored as a “partial loss.” Neurodegeneration resulting from cat-2 overexpression was scored using developmentally synchronized worms, as indicated. A DeltaVision microscope (Applied Precision) was used to acquire images. All images were analyzed using softWoRx Suite and softWoRx Explorer software (Applied Precision).
Softworx suite
Manufactured by Cytiva
Sourced in United States
SoftWoRx Suite is a comprehensive software package designed to support various imaging and analysis capabilities for Cytiva's microscopy systems. The suite provides a range of tools and functionalities to assist users in their research and laboratory workflows.
Automatically generated - may contain errors
Lab products found in correlation
Axiovert 200m, by Zeiss (4 mentions)
Metamorph 6, by Molecular Devices (4 mentions)
Essentials v 3, by Scientific Volume Imaging (3 mentions)
Softworx explorer, by Cytiva (2 mentions)
Coolsnap hq2, by Nikon (2 mentions)
Em c2 emccd, by Cytiva (2 mentions)
Deltavision rt, by Zeiss (2 mentions)
Eclipse ti, by Nikon (2 mentions)
Fluorescent dissecting microscope, by Leica (1 mentions)
Deltavision microscope, by Cytiva (1 mentions)
Deltavision rt system, by Cytiva (1 mentions)
Coolsnap hq charge coupled device camera, by Teledyne (1 mentions)
Ix inverted microscope, by Olympus (1 mentions)
Deltavision system, by Cytiva (1 mentions)
9 protocols using softworx suite
1
Scoring Neurodegeneration in C. elegans
2
Live-cell Imaging Using LabTek Chambers
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Cells were grown in No. 1.5 LabTek II chambered coverglasses (Nalge NUNC, Rochester, NY). To adhere gelatin to the glass, the glass was coated with 0.1% gelatin, air dried, fixed with 4% paraformaldehyde for 2 h, and rinsed thoroughly with PBS. Cells grown using this method had normal morphology compared with standard gelatin-coated plastic dishes. Live-cell imaging was performed using a DeltaVision RT system (Applied Precision, Seattle, WA) with an Olympus IX inverted microscope and a CoolSnap HQ charge-coupled device camera (Roper Scientific, Tucson, AZ). Excitation and emission filters used were as follows: CFP, 436/10 and 465/30 nm; YFP, 492/18 and 535/30 nm; and RFP, 580/20 and 630/60 nm. Images were acquired using a 60× PlanApo 1.4 numerical aperture oil objective (Olympus). An environmental chamber was used at 37°C with 5% CO2 perfusion. Image analysis was performed using SoftWoRx Suite (Applied Precision) and Volocity.
3
Wide-field Fluorescence Microscopy of Bacterial Cells
4
Deconvolution and Analysis of Microscopy Images
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Microscopy images acquired with a Delta Vision microscopy were deconvolved and analysed using softWoRx Suite and softWoRx Explorer software (AppliedPrecision, Issaquah, WA, USA). Images acquired with a spinning-disk confocal microscopy were analyzed by ImageJ software.
5
Wide-field Fluorescence Microscopy Protocol
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Wide-field fluorescence microscopy was performed with cells immobilized on microscope slides covered with a thin film of 1.2% agarose. Unless stated differently, bacterial cell membranes were visualized with Nile Red at a final concentration of 1 μg ml−1. Microscopy was carried out with Zeiss Axiovert 200M (Zeiss Plan-Neofluar × 100/1.30 Oil Ph3 objective, Photometrics CoolSnap HQ2 CCD camera), Nikon Eclipse Ti (Nikon Plan Fluor × 100/1.30 oil Ph3 DLL objective, Rolera EM-C2 EMCCD camera) and Applied Precision DeltaVision RT (Zeiss Plan-Neofluar × 100 Ph3 objective, Photometrics CoolSnap HQ CCD camera). The images were acquired with Metamorph 6 (Molecular Devices) and softWoRx Suite (Applied Precision), and further analysed using ImageJ 1.48 (NIH). Deconvolution was carried from optical sections using Huygens Essentials v.3.3 (Scientific Volume Imaging).
6
Nile Red Lipid Droplet Imaging
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Nile red staining and observation of lipid droplets were as described previously (Hung et al., 2013 (link)) using Nile red (Fluka, Cat. No. 72485) and DeltaVision system (Applied Precision) with a 100x objective lens (NA = 1.4) and a CoolSNAP HQ CCD camera (Photomotrics) controlled by softWoRx Suite (Applied Precision). Normarski differential interference contrast (DIC) microscopy image was taken as previously described (Fu et al., 2010 (link)).
7
Fluorescence Microscopy of Cellular Membranes
8
Fluorescence Microscopy of Membrane Potential
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For fluorescence microscopy, cells were grown to exponential growth phase at 30 °C if not stated otherwise. The cells were immobilized on microscope slides covered with a thin film of 1.2% agarose in water. For the dissipation of membrane potential with CCCP, the mounting medium was supplemented with 0.5% dimethylsulphoxide (DMSO) or 100 μM CCCP dissolved in DMSO (0.5% final concentration of DMSO). Imaging was carried out within 2 min after addition of ionophores. Membranes were visualized with Nile Red or FM 4-64 (0.5 μg ml−1). Standard fluorescence microscopy was carried out using Zeiss Axiovert 200 M (Zeiss Plan-Neofluar × 100/1.30 Oil Ph3 objective), Nikon Eclipse Ti (Nikon Plan Fluor × 100/1.30 Oil Ph3 DLL objective), and Applied Precision DeltaVision RT (Zeiss Plan-Neofluar 100x/1.30 Oil Ph3) microscopes. The images were acquired with Metamorph 6 (Molecular Devices), softWoRx Suite (Applied Precision), and further analysed using ImageJ v.1.38 (National Institutes of Health). Deconvolution was carried from optical sections using Huygens Essentials v.3.3 (Scientific Volume Imaging).
9
Imaging Vacuole Microdomains and Lipid Droplet Dynamics
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Cells for microscopy were either prepared in SC medium or in 50 mM Tris-HCl, pH 7.5. The images were acquired at 30oC by an imaging system (DeltaVision; Applied Precision) with a Plan Apochromat 100× objective lens, NA 1.4, and a charge-coupled device camera (CoolSNAP HQ2; Photometrics) controlled by softWoRx suite (Applied Precision). The filters used with the DeltaVision system were GFP (525/50 nm), mCherry (632/60 nm) YFP (559/38 nm), and CFP (470/24 nm). To image the microdomains of vacuoles, cells were imaged from the periphery by taking nine optical section images spaced at 0.3 µm. Images were further processed by deconvolution and maximal projection by softWoRx suite. To image LDs entering vacuoles by 4D (3D time lapse), we used the DeltaVision system equipped with a 512 × 512–pixel electron-multiplying charge-coupled device camera (Evolve 512; Photometrics) or CoolSNAP HQ2 charge-coupled device camera controlled by softWoRx suite.
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