After euthanasia at 4 and 8 weeks, the knee joins were fixed with 4% PFA and decalcified in ethylene diamine tetra acetic acid (EDTA, pH = 7.2, Macklin, China) by Ultrasonic Decalcifying Unit (USE 33, Germany) for 1 month. The joint samples were embedded and sectioned in paraffin (5-μm thickness) for further staining. The joint sections were respectively stained with H&E (Solarbio, China) and Safranine O-fast green (Safranine O, Solarbio, China) for histological analysis, and OA progression was evaluated according to Mankin scoring as previously described by 2 blinded observers [41 (link)]. Specifically, for immunohistochemical staining, the sections were initially incubated with rabbit polyclonal anti-IL6 or anti-MMP13 (Boster, China) antibodies overnight at 4 °C followed by binding with biotinylated secondary antibodies (ZS-Bio, China). The slides were photographed by optical microscopy (OLYMPUS BX53F, Japan).
Safranine o fast green
Manufactured by Solarbio
Sourced in China
Safranine O-Fast Green is a dual stain solution used for staining biological samples in laboratory settings. It consists of two dyes, Safranine O and Fast Green FCF, which can be used to differentiate between various cellular structures and components.
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Lab products found in correlation
3 protocols using safranine o fast green
1
Histological Analysis of Osteoarthritis Progression
2
Histological Assessment of Rat Ankle Joint
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Overdose of sodium pentobarbital euthanized the rats. The rat ankle joint samples were fixed in cold paraformaldehyde solution (4%) for 48 h, and then decalcified in 0.5 M EDTA solution for 70 days. Then, the ankle joint was embedded in paraffin and a paraffin section (5-μm thick) was used for histological analysis. Haematoxylin-Eosin (H&E) staining kit (Beyotime, China), Safranine O-Fast Green (Solarbio, China), and TRAP (Sigma, USA) staining were performed according to standard protocols, respectively. After staining, we observed the pathological changes in tissues under an optical microscope. H&E scored [16 (link)] from 0 to 4; 0, normal joint synovial tissue and bone structure; 1, synovial cell hypertrophy and inflammatory cell erosion of synovial tissue; 2, cartilage destruction and pannus present; 3, most articular cartilage and subchondral bone are destroyed; 4, joint adhesions and stiffness and accompanying disability. Toluidine Blue scored [17 (link)] from 0 to 3, where: 0, normal; 1, slight cartilage erosion; 2, moderate cartilage erosion; 3, severe cartilage erosion or bone destruction. The number of TRAP-positive cells was quantified.
3
Histopathological Evaluation of Ankle Joint
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Ankle joints were fixed for 24 h in 4% buffered formalin (Sigma–Aldrich). The formalin was washed with water, and the tissue was decalcified (Sigma–Aldrich) and embedded in paraffin. Ankle joints tissues were then sliced into 5 μm thick sections. H&E (Beyotime, Beijing, China) and Safranine O-Fast Green (Solarbio, Beijing, China) staining were performed according to standard protocols. After staining, observe the changes of pathological tissues under an optical microscope. H&E scored [18 (link)] from 0 to 4, 0 = normal joint synovial tissue and bone structure; 1 = synovial cell hypertrophy and inflammatory cell erosion of synovial tissue; 2 = cartilage destruction and pannus present; 3 = most articular cartilage and subchondral bone are destroyed; 4 = joint adhesions and stiffness and accompanying disability. Safranine O-Fast Green staining was scored [19 ] from 0 to 3, where 0 = normal, 1 = slight cartilage erosion, 2 = moderate cartilage erosion, and 3 = cartilage erosion or bone destruction.
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