Mice were euthanized by exposure to gradually increasing concentrations of carbon dioxide. Dissected TA and quadriceps were stuck onto the pedestal by tragacanth gum (Wako, 206-02242) on a cork and frozen by dipping into 2-methylbutane (Wako, 166-00615) for 45–60 s at melting point temperature (−156°C). Freshly isolated diaphragms were embedded in OCT compound (Sakura Finetek, 4583, USA) and directly frozen within 2-methylbutane. Frozen muscles were sliced at 12-μm thickness with a Cryostat (Leica, CM1850) and attached to the slide. The sample on the slide was kept at −80°C for the following immunostaining.
Tragacanth gum
Manufactured by Fujifilm
Sourced in Japan
Tragacanth gum is a natural, water-soluble polysaccharide obtained from the dried sap of certain species of the Astragalus plant. It is commonly used as a thickening, stabilizing, and emulsifying agent in various laboratory and industrial applications.
Automatically generated - may contain errors
Lab products found in correlation
2 methylbutane, by Sakura Finetek (1 mentions)
2 methylbutane, by Fujifilm (1 mentions)
Oct compound, by Sakura Finetek (1 mentions)
2 methylbutane, by Merck Group (1 mentions)
Lsm 710 confocal laser scanning microscope, by Zeiss (1 mentions)
Hybrid cell count software bz h3c, by Keyence (1 mentions)
Bx51 microscope, by Olympus (1 mentions)
Protein block, by Agilent Technologies (1 mentions)
Alexa fluor 594, by Abcam (1 mentions)
Ncl dys2, by Leica Biosystems (1 mentions)
Bz x710, by Keyence (1 mentions)
Isopentane, by Fujifilm (1 mentions)
Entellan new, by Merck Group (1 mentions)
Alexa 555 conjugated phalloidin, by Thermo Fisher Scientific (1 mentions)
Cryostat, by Leica camera (1 mentions)
Paraformaldehyde (pfa), by Merck Group (1 mentions)
Hoechst 33342, by Merck Group (1 mentions)
Bz 9000, by Keyence (1 mentions)
6 protocols using tragacanth gum
1
Cryopreservation of Mouse Muscle Tissues
2
Muscle Tissue Cryopreservation and Histochemical Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Before examination, the fish was anesthetized by 0.025% buffered MS-222 solution (Sigma-Aldrich, MI, USA) following a previously reported method [25 (link)]. We cut the muscle tissue between the anal and caudal fins and placed the muscle tissue in wet gauze. We placed 10% Tragacanth gum (FUJIFILM Wako Pure Chemical Corporation, Tokyo, Japan) on the well, which was stirred evenly, and formed into a small hill shape. We placed the muscle tissue into the small hill. Then, a beaker filled with 100 mL 2-methylbutane (Sigma-Aldrich, MI, USA) and placed into liquid nitrogen. We completely submerged the wood with muscle tissue into 2-methylbutane for 1 min. We then transferred muscle tissue to a − 80 °C freezer immediately. The muscle tissue was sectioned into 8 µm slices using Cryostat Microtome CM3050S (Leica, Wetzlar, Germany). Tissue sections were stored in a − 80 °C freezer, and later stained with Hematoxylin and Eosin stain (HE stain), Gomori’s trichrome stain, and nicotinamide adenine dinucleotide dehydrogenase-tetrazolium reductase stain (NADH-TR stain).
3
Oral Zonisamide Administration in Viral Study
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ZNS was provided by Dainippon Sumitomo Pharma. ZNS was suspended in a 0.5% tragacanth gum (Wako) solution before use. We administrated the ZNS solution at a dose of 40 mg/kg by oral gavage once daily (ZNS group). We continued the administration of ZNS until the day before sacrifice as described later. As a control, the same volume of vehicle was simultaneously administrated in a separate set of subjects (vehicle group). Unless otherwise stated, ZNS administration was started 3 days before the viral injection.
4
Muscle Regeneration Histological Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mice were sacrificed 3 days, 2 weeks, and 5 weeks post injury. TA muscles were mounted in Tragacanth Gum (FUJIFILM Wako) and frozen with liquid nitrogen47 (link). Serial sections (10 µm) were cut using a cryostat. Sections were stained with H&E and observed using an Olympus BX51 microscope (Olympus, Tokyo, Japan). Four sections prepared from the middle of each TA muscle samples were stained, and the entire transverse section of all sections was photographed and analyzed. Of the values obtained from the four thin sections taken, the one with the highest value was adopted as the data for each sample. Immunofluorescence images were acquired using the Zeiss LSM 710 laser scanning confocal microscope (Zeiss). Area measurement and cell counting were performed using Hybrid cell count software BZ-H3C (Keyence). In Fig. 6C , images stained with Laminin antibody were analyzed with KEYENCE image analysis software to calculate the average area of myofibers. In Fig. 6E , images stained with MYH4 antibody were analyzed with KEYENCE image analysis software to calculate the area of the positive area. In Fig. 6G , images stained with MYH3 antibody were counted for the number of positive fibers.
5
Dystrophin Expression in DYS-HAC Pigs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After DYS-HAC-cloned pigs were euthanized under general anesthesia, the skeletal muscles (biceps femoris) were dissected, mounted on cork bases using tragacanth gum (Fujifilm Wako Pure Chemical, Osaka, Japan), and frozen by isopentane cooled in liquid nitrogen. Histological analyses were performed on 8-μm-thick frozen sections. After the observation of EGFP expression in the frozen section under fluorescence microscopy (BZ-X710, Keyence, Osaka, Japan), immunofluorescence staining for dystrophin was performed using the same frozen section. The histological serial sections were stained with H&E using a standard technique. For immunofluorescence, the sections were incubated with Protein Block (X0909, Dako, Glostrup, Denmark) for 30 min at 25°C and then treated with Mouse Monoclonal Antibody Dystrophin (NCL-DYS2, 1:50 dilution, Leica Biosystems, Wetzlar, Germany), which reacts with both human and pig dystrophin, for 1 h at room temperature. After the removal of excess antibody, the sections were incubated with Alexa Fluor 594 (ab150112,1:450 dilution, Abcam, Cambridge, UK) for 1 h at room temperature. The slides were visualized under fluorescence microscopy (BZ-X710, Keyence).
6
Fluorescent Imaging of Skeletal Muscle Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Muscle tissue isolated from the left hind limb was stuck to a corkboard piece using Tragacanth gum (Wako). Then, the skeletal muscle was rapidly frozen in isopentane (Wako) pre-chilled in liquid nitrogen and stored at −80 °C until sectioning. Seven µm-thick cross-sections were made from muscle samples with a cryostat (Leica, Wetzlar, Germany) at −20 °C, fixed with 4% paraformaldehyde (Sigma-Aldrich) in PBS, and then stained with Alexa555-conjugated phalloidin (Thermo Fisher Scientific, Waltham, MA, USA) and Hoechst 33342 (Sigma-Aldrich). After staining, the cross-sections were washed with PBS and mounted using Entellan new (MERCK, Darmstadt, Germany). Stained muscle sections were observed on a fluorescence microscope (BZ-9000: KEYENCE, Osaka, Japan), and captured fluorescent images of the cross-sections were analyzed using the ImageJ software (Fiji package; NIH, Bethesda, MD, USA).
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!