Caudal fin fold amputation was performed in 3 dpf larvae under anaesthesia with 0.016% Tricaine (MS222, Sigma) in zebrafish water using a sterile scalpel57 (link). For imaging, live embryos were anesthetized in 0.016% Tricaine, and positioned in 35 mm glass-bottom dishes (FluoroDishTM, World Precision Instruments). They were mounted in 1% low melting point agarose (Sigma) with Tricaine. Light microscopy was performed using an MVX10 Olympus macroscope equipped with an MVPLAPO 1X objective and XC50 camera. Z-stacks series were obtained using an inverted confocal microscope Leica TCS SP5 (Leica Application Suite V3.2) and TCS SP8 (Leica Application Suite V3.5) equipped with an HCXPL APO 40x/1.25-0.75 oil and an HC PL APO 0.70 ∞ (infinity) 20x objective (Leica). The mCherry signal was excited with a 560 nm laser, and GFP with a 490 nm laser. Datasets were analysed using Fiji Software (ImageJ 1.52p)58 (link).
Fluorodishtm
Manufactured by World Precision Instruments
Sourced in Germany, United States
The FluoroDishTM is a specialized petri dish designed for fluorescence microscopy applications. It features a high-quality glass bottom that provides optimal optical clarity and performance for fluorescent imaging. The dish is constructed using durable materials to provide a reliable platform for cell culture and various other life science experiments.
Automatically generated - may contain errors
Lab products found in correlation
Ms 222, by Merck Group (1 mentions)
Mvplapo 1x objective, by Olympus (1 mentions)
Mvx10 macroscope, by Olympus (1 mentions)
Leica tcs sp5, by Leica camera (1 mentions)
Tcs sp8, by Leica camera (1 mentions)
3d cell explorer, by Nanolive (1 mentions)
Rg208007, by OriGene (1 mentions)
Ps100010, by OriGene (1 mentions)
Eclipse te2000 u microscope, by Nikon (1 mentions)
Fetal bovine serum (fbs), by ScienCell (1 mentions)
Huvecs, by Lonza (1 mentions)
Endothelial cell medium (ecm), by ScienCell (1 mentions)
Endothelial cell growth supplement (ecgs), by ScienCell (1 mentions)
5 protocols using fluorodishtm
1
Caudal Fin Fold Amputation in Zebrafish
2
Murine Sperm Demembranation and Reactivation
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In a petri dish, murine sperm of the stock solution (10 µL) were demembranated and reactivated in 3 mL of a demembranation and reactivation buffer (pH 7.8, in mM: 132 sucrose, 24 potassium glutamate, 20 Tris, 0.1% Triton X-100, 1 Dithiothreitol (DTT), 1 MgCl2, 0.5 EGTA, 1 ATP and 3 µM cAMP). The necessary amounts of ATP, ADP, cAMP and Mg2+ were added to the demembranation/reactivation buffer. The reactivated sperm were transferred to a FluoroDishTM (World Precision Instruments, Friedberg, Germany) for local microperfusion or transferred to 100 µm deep chamber slides (Leja, Nieuw-Vennep, The Netherlands) for holographic imaging. All experiments were performed at room temperature.
3
Imaging human astrocyte NOX4 expression
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Human astrocytes (2 × 104 cells) were seeded in FluoroDish TM (FD35-100, World Precision Instruments, Sarasota, FL, USA). Cells were transduced with pCMV6-AC-GFP constructs against human NOX4 (NM_016931) (RG208007, Origene, Rockville, MD, USA) or pCMV6-AC-GFP vector (PS100010, Origene). Cells were incubated for 24 h or 48 h 3D images were analyzed with 3D Cell Explorer (NANOLIVE, Ecublens, Switzerland). Images were representative images from a total of 100 cells in ten individual images per group.
4
Flagellar Waveform Analysis in Sperm
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The flagellar waveform was analyzed as previously described [30 (link)] with a Nikon Eclipse TE2000-U microscope. Images were collected at 300 Hz respectively by a M3 high speed camera (IDT, Tallahassee, FL, USA) using a 20× objective and the Motion-Studio 64 software (Imaging Solutions, Ehingen, Germany). For sperm beat frequency analysis, 10 µL of the sperm suspension in demembranation/reactivation buffer was transferred to a FluoroDishTM (World Precision Instruments, Germany) and were allowed to adhere to the bottom during 3 Min incubation. Cells were perfused with different concentrations of ATP, ADP and Mg2+ for one minute (Figure 1 B). Single sperm were selected for analysis with Image J (1.48V, National Institute of Health, Washington, DC, USA). Flagellar beat frequency was measured with semi-automated Igor ProTM software (Wavemetrics, Lake Oswego, OR, USA) like previously described [30 (link),68 (link)].
5
Culturing HUVECs on Fibronectin-Coated PDLLA Nanofilms
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Human Umbilical Vein Endothelial Cells (HUVECs) (Lonza, Cat. # 00191027), chosen as a model for endothelial cells, were cultured in Endothelial Cell Medium (ECM, ScienCell TM , USA, Catalog 1001) supplemented with 1% Endothelial Cell Growth Supplement (ECGS, ScienCell TM , USA, Catalog 1052), 5% Fetal Bovine Serum (FBS, ScienCell TM , USA, Catalog 0025), 1% Pen-Strep mixture (ScienCell TM , USA, Catalog 0503) and stored in an incubator at 37°C and 5%CO2. HUVECs were sub-cultured in conventional T75 flasks up to passage 8. For the experiment, cells were plated at a concentration of ~10,000 cells/cm 2 onto 9 porous PDLLA nanofilms adherent to cell culture Petri dishes. Glass bottom Petri dishes were used as control (FluoroDish TM , World Precision Instruments, Inc.,USA, Cat.# FD35-100). Before culturing cells on PDLLA, the nanofilm was coated with bovine fibronectin (FN, Sigma-Aldrich Co. LLC., St. Louis, MO) aqueous solution: nanofilms were exposed to oxygen plasma (3 min at 200 mTorr and 18 W), then FN solution was deposited at the density of 5 µg/cm 2 .
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