To analyze catabolic activity in the alveolar bone, multinucleated osteoclasts and preosteoclasts were stained with tartrate-resistant acid phosphatase (TRAP). After deparaffinization and rehydration, the sections were stained using a TRAP staining kit (Wako Pure Chemical, Osaka, Japan) according to the manufacturer’s protocol. Incubation of the sections in TRAP buffer for 30 min was performed, followed by washing with distilled water, and counterstained with 0.02% Fast Green (Wako Pure Chemical, Osaka, Japan) nuclear staining solution for 10 min. The number of TRAP-positive multinucleated osteoclasts in the PDL area around the M2 mesial roots was counted by a single examiner in three randomized sections for each sample and the averages were calculated.
Fast green
Manufactured by Fujifilm
Sourced in Japan
Fast Green is a laboratory dye used for staining and visualizing proteins in analytical techniques such as gel electrophoresis and Western blotting. It is a water-soluble, green-colored dye that binds to proteins, allowing for their detection and quantification.
Automatically generated - may contain errors
Lab products found in correlation
Trap staining kit, by Fujifilm (1 mentions)
Safranin o, by Merck Group (1 mentions)
Rm2135, by Leica (1 mentions)
Aps coated glass slides, by Matsunami (1 mentions)
Technovit 3040, by Kulzer (1 mentions)
Technovit 7100 resin, by Kulzer (1 mentions)
Dmi8 microscope, by Leica (1 mentions)
Mildform, by Fujifilm (1 mentions)
Sirius red solution, by Fujifilm (1 mentions)
Las x version 3, by Leica (1 mentions)
Mildform 10n, by Fujifilm (1 mentions)
Nanozoomer s60, by Hamamatsu Photonics (1 mentions)
Haematoxylin and eosin, by Merck Group (1 mentions)
Rat anti mouse f4 80 monoclonal antibody, by Bio-Rad (1 mentions)
Peroxidase conjugated streptavidin, by Agilent Technologies (1 mentions)
Dp73 digital camera, by Olympus (1 mentions)
Sirius red, by Merck Group (1 mentions)
Xylene, by Fujifilm (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Fujifilm (1 mentions)
Ethanol, by Fujifilm (1 mentions)
7 protocols using fast green
1
Osteoclast Quantification in Alveolar Bone
2
Histological Analysis of Haustorium Development
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Field-collected samples were washed with tap water to remove soil. Safranin-O staining of whole haustoria was performed as previously described (Yoshida and Shirasu 2009 (link)). For histological sections, haustoria and infected host roots were separated and fixed with FAA solution (10% formaldehyde, 5% acetic acid, 50% ethanol) under vacuum for 15 min followed by incubation at room temperature for 2 h. Samples were subjected to an ethanol dehydration series and embedded into Technovit 7100 resin (Heraeus Kulzer GmbH) according to the manufacturer’s instruction. Solidified resin blocks were mounted on wood blocks with Technovit 3040 (Heraeus Kulzer GmbH), then sectioned using a microtome (Leica, RM2135) to 4 μm thickness. Sections were stained with 0.1% Safranin-O (Sigma-Aldrich) and counterstained with 0.1% Fast Green (Wako Chemical) on APS-coated glass slides (Matsunami Glass).
3
Histological Assessment of Liver Pathology
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The mice liver tissues were fixed in Mildform (Fujifilm Wako Chemicals, Osaka, Japan), processed for paraffin embedding, and sectioned into 3 µm-thick sections. Each section was deparaffinized, rehydrated, and processed for hematoxylin–eosin and Sirius red-Fast Green staining. Sections were incubated in 0.04% Fast Green (Fujifilm Wako Chemicals), saturated in picric acid for 5 min, washed with distilled water, and incubated in 0.1% Sirius Red solution (Fujifilm Wako Chemicals) for 10 min. The histological features after each staining were determined using a Leica DMi8 microscope and LAS X version 3.3 (Leica Microsystems, Inc., Wetzlar, Germany). The steatosis, inflammation, and fibrosis degrees were assessed by an experienced pathologist blinded to the experimental procedures according to previously reported criteria33 (link),34 (link).
4
Modulating LIF Signaling in Fetal Cerebrum
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According to our previous study, the serum level of LIF reached its peak at E14.5 in pregnant dams of rats, and maternally injected LIF induced NPCs proliferation in the fetal cerebrum in rats [14 (link)]. Therefore, we injected 5 μg/kg body weight (BW) recombinant rat LIF (3010, MilliporeSigma, Burlington, MA, USA) into pregnant dams intraperitoneally at E14.5 to analyze total mRNA expression using DNA microarray and Igf1 and Igf2 expression using quantitative RT-PCR. In addition, we also examined the effects of LIF directly injected into the lateral ventricle of the fetal cerebrum at E14.5 via in utero injection technique [38 (link)]. For in utero injection, 1 μL of recombinant rat LIF (5 ng/μL) diluted with PBS and Fast Green (100:1, FUJIFILM Wako Pure Chemical, Osaka, Japan) was administered to each fetus. The same volume of PBS was injected into the lateral ventricle of control fetuses. Three hours after the injection, the dorsal cerebrum was collected and stored as a pooled sample for each group.
5
Histological Analysis of c-Fos KO Rat Cartilage
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Under anesthesia with 3.5% isoflurane inhalation, 4-week-old c-Fos KO rats were perfused with saline, followed by Mildform 10N (131-10317; FUJIFILM Wako Pure Chemical,
Osaka, Japan). Tissue samples were decalcified in 10% EDTA (pH 7) for 1 week. Decalcified tissues were embedded in paraffin. Paraffin‐embedded tissues were sliced into
5-µm-thick sections and analyzed using hematoxylin and eosin (H&E) staining. Safranin O (Waldeck GmbH & Co., Havixbecker, KG)‐Fast Green (FUJIFILM Wako Pure
Chemical) staining was performed to observe the morphology of cartilage tissues. The images were captured by Nanozoomer S60 (Hamamatsu Photonics, Hamamatsu, Japan). The quantification of the
epiphysis and medullary cavity area with the images was performed using ImageJ 1.53k software (National Institutes of Health, Bethesda, MD, USA).
Osaka, Japan). Tissue samples were decalcified in 10% EDTA (pH 7) for 1 week. Decalcified tissues were embedded in paraffin. Paraffin‐embedded tissues were sliced into
5-µm-thick sections and analyzed using hematoxylin and eosin (H&E) staining. Safranin O (Waldeck GmbH & Co., Havixbecker, KG)‐Fast Green (FUJIFILM Wako Pure
Chemical) staining was performed to observe the morphology of cartilage tissues. The images were captured by Nanozoomer S60 (Hamamatsu Photonics, Hamamatsu, Japan). The quantification of the
epiphysis and medullary cavity area with the images was performed using ImageJ 1.53k software (National Institutes of Health, Bethesda, MD, USA).
6
Histological Analysis of Liver Fibrosis
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Liver tissue was fixed in 10 % (v/v) neutral buffered formalin solution, dehydrated with ethanol, cleared in xylene and embedded in paraffin. Then the paraffin-embedded blocks were cut into sections approximately 5 µm thick. After removal of paraffin with xylene, sections were stained with haematoxylin and eosin (Merck)(30 ) for morphological analysis, or were stained with Sirius red (Sigma-Aldrich) and counterstained with fast green (Wako) for determination of the area of fibrosis(31 (link)). Immunohistochemical staining of hepatic monocytes/macrophages was performed by using sections of formalin-fixed, paraffin-embedded liver tissue as described previously(32 (link),33 (link)). Briefly, after removal of paraffin, sections were incubated with a rat anti-mouse F4/80 monoclonal antibody (Serotec), followed by incubation with a biotinylated rabbit anti-rat IgG antibody (Dako) and peroxidase-conjugated streptavidin (Dako). Then colour was developed with diaminobenzidine tetrahydrochloride (Dojindo) and the sections were counterstained with haematoxylin. Images were acquired with an Olympus DP73 digital camera (Olympus IX-73; Olympus) under an inverted microscope (original magnification, ×140). The F4/80-positive area and Sirius red-positive area were quantified as a percentage of the total tissue area by using cellSens Dimension Olympus 1.15 software (Olympus).
7
Histological and Immunohistochemical Analysis
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Paraffin-embedded tissue sections were deparaffinized using xylene (Fujifilm Wako, Osaka, Japan) and ethanol (Fujifilm Wako) and stained with hematoxylin (Sigma) & eosin (Sigma), Safranin O (Nacalai Tesque, Kyoto, Japan), and Fast Green (Fujifilm Wako). For immunostaining, we incubated the tissue sections for 40 min at 37 • C in 10 mg/mL hyaluronidase (Sigma) and 15 min at 80 • C in 1 mM EDTA (Fujifilm Wako, Osaka, Japan) for antigen retrieval. Blocking was performed in 10% FBS (Nichirei, Tokyo, Japan) for 1 hour, and the sections were incubated at 4 • C overnight with antibodies against COL 1 (Southern Biotechnology Associates, Birmingham, AL, #1310-01) at 1:500 and COL 2 (Southern Biotechnology Associates, #1320-01) at 1:600, and COL 10 (Thermo Fisher Scientific, #14-9771-80) at 1:300. On the following day, the sections were stained with Simple Stain MAX-PO (G) (Nichirei) to detect the signals for COL 1 and COL 2 and with Alexa 555-conjugated goat anti-mouse IgG antibody (Thermo Fisher Scientific, #A21425) for COL 10. The nuclei were stained with 10 µg/mL DAPI (R&D, Minneapolis, MN), after which the slides were observed with an All-in-One Fluorescence Microscope BZ-X810 (Keyence, Osaka, Japan).
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