Mono- and dual-species biofilms were stained with 5 μM calcofluor white (Invitrogen, Paisley, UK) and 20 μM SYTO9® (Sigma–Aldrich, Dorset, UK) was used to stain both fungal and bacterial cells. Biofilms were then imaged using a confocal laser scanning microscopy (CLSM) microscope (Leica SP5 laser scanning confocal microscope) at an excitation and emission wavelength of 350 and 400 nm for calcofluor white and 485 and 500 nm for SYTO9®. Images were then processed and analyzed using Volocity 3D Image Analysis Software (Perkin Elmer). In addition, biofilms were also imaged using scanning electron microscopy (SEM). After biofilm development, biofilms were fixed using 2% para-formaldehyde, 2% glutaraldehyde, 0.15 M sodium cacodylate, and 0.15% w/v alcian blue and processed for SEM, as previously described (Erlandsen et al., 2004 (link)). Samples were then sputter coated in gold before being imaged using a JEOL JSM-6400 scanning electron microscope.
Calcofluor white
Manufactured by Leica
Sourced in Germany
Calcofluor white is a fluorescent stain used in microscopy to visualize and detect the presence of cellulose and chitin in biological samples. It binds to these polysaccharides, emitting a bright blue fluorescence under ultraviolet or blue light excitation.
Automatically generated - may contain errors
Lab products found in correlation
Syto9, by Merck Group (1 mentions)
Calcofluor white, by Thermo Fisher Scientific (1 mentions)
Volocity 3d image analysis software, by PerkinElmer (1 mentions)
Jsm 6400 scanning electron microscope, by JEOL (1 mentions)
Sp5 laser scanning confocal microscope, by Leica (1 mentions)
Syto9, by Leica (1 mentions)
Dm2500, by Leica (1 mentions)
Szx16, by Olympus (1 mentions)
Calcofluor white solution, by Merck Group (1 mentions)
Syto13, by Thermo Fisher Scientific (1 mentions)
Dfc425c camera, by Leica (1 mentions)
Tcs sp8, by Leica (1 mentions)
Toluidine blue o, by Merck Group (1 mentions)
Toluidine blue, by Merck Group (1 mentions)
Calcofluor white, by Merck Group (1 mentions)
Dm5500b microscope, by Leica (1 mentions)
Rj2035 microtome, by Leica (1 mentions)
Lr white, by Electron Microscopy Sciences (1 mentions)
3 protocols using calcofluor white
1
Multi-Modal Imaging of Mono- and Dual-Species Biofilms
2
Seed Cell Wall Staining Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The staining procedure was performed as previously described [24 (link)], with slight modifications, as follows. After neutralization of potassium hydroxide (KOH) treatment, the seeds were stained with 40 µg/mL WGA-Alexa fluor-488 (Alexa-488) solution (Thermo Fisher Scientific, Waltham, MA, USA) for 15 min and 20 µg/mL Calcofluor White solution (Sigma-Aldrich, St. Louis, MO, USA) for 5 min in the dark at room temperature. A seed coat of the stained intact seed was removed under a stereomicroscope (SZX16; Olympus, Tokyo, Japan). Stained cells were observed under a fluorescence microscope (DM2500; Leica, Wetzlar, Germany) with an excitation filter of 480/40 nm for Alexa-488-conjugated fungal hyphae and 340–380 nm for Calcofluor White-stained plant cell walls.
3
Microscopic Analysis of Peanut Root Nodules
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For a macroscopic view, pictures were taken under a Nikon SMZ25 stereomicroscope and images were processed using NIS Elements AR 5.10.00. For semithin sectioning, peanut root nodules were fixed at 4°C overnight with 3% paraformaldehyde (wt/vol) and 2.5% glutaraldehyde (vol/vol) in 0.05 M sodium cacodylate buffer (pH 7.2). The fixed samples were dehydrated in ethanol gradient and subsequently embedded in LR white (Electron Microscopy Science), and semithin sections were generated according to the manufacturer's protocol. Sections (1 μm) were cut with an RJ2035 microtome (Leica Microsystems). For Toluidine Blue staining, the sections were stained for 5 min in 0.05% Toluidine Blue O (Sigma) and analyzed using a DM5500B microscope equipped with a DFC425C camera (Leica Microsystems). For confocal microscopy, sections were cut using a vibratome (Leica Microsystems, VT-1000S) and stained with Calcofluor white (Sigma) and SYTO13 (Life Technologies). Confocal images were captured with a TCS-SP8 (Leica) using excitation (ex) of 488 nm and emission (em) of 500 to 530 nm for SYTO13 and 405 nm ex and 410 to 430 nm em for Calcofluor white. Images were processed with LAS AF Lite version software and prepared for presentation with Photoshop CS6 (Adobe Systems Inc.).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!