The WT and BTN1A1(−/−) BMEC were plated at 60,000 cells per 35-mm Glass Bottom Cell Culture Dish (NEST Biotechnology, China) for oil red staining (n = 3). After culturing with lactogenic medium for 48 h, Oil-Red-O staining was performed using Oil-Red-O working solution (G1262, Solarbio) with modifications. Briefly, cells were washed thrice with PBS and fixed with 4% paraformaldehyde for 20 min at room temperature. After three washes in PBS, cells were washed with isopropanol and stained with Oil-Red-O working solution (G1260, Solarbio) for 25 min, then washed with PBS, and nuclei re-stained with hematoxylin for 1–2 min. Cells were covered with distilled water and images were visualised using an Olympus IX73 fluorescence microscope equipped with an Olympus DP80 digital camera. Forty LD were randomly selected from each cell culture dish and diameter measured using Cell Sens standard software (version 1.13; Olympus). Average LD diameter values represent the mean size of all 120 LD per group. Lipid droplets were divided into four size groups (0 < size < 2.0 μm, 2.0 μm < size < 2.5 μm, 2.5 μm < size < 3.0 μm and size > 3.0 μm). The number of LD were counted within each of these four sizes.
G1260
Manufactured by Solarbio
Sourced in China
The G1260 is a laboratory equipment designed for general use. It serves as a general-purpose tool for various applications in a laboratory setting.
Automatically generated - may contain errors
Lab products found in correlation
G1262, by Solarbio (2 mentions)
Oil red o working solution, by Solarbio (1 mentions)
Ix73 fluorescence microscope, by Olympus (1 mentions)
Glass bottom cell culture dish, by NEST Biotechnology (1 mentions)
Cellsens standard software, by Olympus (1 mentions)
Mesencult acf chondrogenic differentiation medium, by STEMCELL (1 mentions)
D1756, by Merck Group (1 mentions)
G1452, by Solarbio (1 mentions)
Dexamethasone (dex), by Merck Group (1 mentions)
Indomethacin, by Merck Group (1 mentions)
β glycerophosphate, by Merck Group (1 mentions)
Toluidine blue, by Solarbio (1 mentions)
Sirius red, by Solarbio (1 mentions)
G1285, by Solarbio (1 mentions)
Ab pas, by Solarbio (1 mentions)
Masson g1340, by Solarbio (1 mentions)
Masson trichrome, by Solarbio (1 mentions)
G1120 for h e, by Solarbio (1 mentions)
6 protocols using g1260
1
Oil Red O Staining of Lipid Droplets
2
Multilineage Differentiation Assay for CSPCs
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For chondrogenesis, 1 × 106 CSPCs at passage 2 were centrifuged in 15 mL polypropylene tubes to obtain cell pellets and cultured in MesenCult™-ACF chondrogenic differentiation medium (Catalogue #05455, Stem Cell) for 4 weeks. After differentiation, pellets were fixed and sectioned. Chondrogenesis was analysed by staining with toluidine blue (G3668, Solarbio). For osteogenesis, CSPCs at passage 2 were harvested and cultured in 24-well plates (20,000 cells/well). After 90% confluence, the osteogenic differentiation medium was replaced [low glucose DMEM (10567-014, Gibco) with 10% FBS, 10 mM β-glycerophosphate (50020, Sigma), 50 mM L-ascorbate-2-monophosphate (A7631, Sigma), and 1 μM dexamethasone (D1756, Sigma)] for 3 weeks. The osteogenic cells were fixed and stained with alizarin red (G1452, Solarbio). To induce adipogenesis, CSPCs at passage 2 were harvested and cultured in 24-well plates (20,000 cells/well). After 90% confluence, the osteogenic differentiation medium high glucose DMEM (11965-092, Gibco) was replaced with 10% FBS, 100 μM indomethacin (I7378, Sigma), 0.5 mM IBMX (I5879, Sigma), 10 μg/mL insulin (91077C, Sigma), and 1 μM dexamethasone (D1756, Sigma) for 2 weeks. Oil Red O (G1260, Solarbio) staining was performed to assess the adipogenicity potency.
3
Histological Evaluation of Liver Samples
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Samples were fixed in 4% paraformaldehyde solution and embedded in paraffin. Then, 5 μm sections were made and mounted on slides for staining with hematoxylin and eosin (H&E, G1120, Solarbio, Beijing, China), Masson (G1340, Solarbio, Beijing, China), Sirius Red (G3632, Solarbio, Beijing, China) and AB-PAS (G1285, Solarbio, Beijing, China), according to the manufactures’ instructions. For Oil Red O staining, frozen sections of the livers were first obtained, and the staining was performed according to the manufacturer’s instructions (G1260, Solarbio, Beijing, China), while the quantification was performed according to a previous protocol [91 (link)].
With H&E staining, hepatocellular steatosis was graded from 0 to 3 based on the percentage of hepatocytes involved (0 = <5%; 1 = 5–33%; 2 = 33–66%; 3 = >66%), according to a previous study [92 (link)]. The NAS score was calculated by steatosis (0–3), lobular inflammation (0−3) and ballooning (0−2), also according to a previous study [32 (link)].
With H&E staining, hepatocellular steatosis was graded from 0 to 3 based on the percentage of hepatocytes involved (0 = <5%; 1 = 5–33%; 2 = 33–66%; 3 = >66%), according to a previous study [92 (link)]. The NAS score was calculated by steatosis (0–3), lobular inflammation (0−3) and ballooning (0−2), also according to a previous study [32 (link)].
4
ORO Staining for Myelin Debris Detection
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ORO staining was performed to detect the accumulation of myelin debris in the LC after SCI and the accumulation of myelin debris in RAW 264.7 cells. For cell experiments, RAW 264.7 cells were treated with myelin debris or D-4F, and the concentration gradient settings of the two were based on previous studies [17 (link)]. The cells to be treated were first washed twice with PBS and fixed with an ORO fixative (G1262, Solarbio, China) for 20–30 min. The fixative was discarded, and the cells were washed twice with distilled water and stained with ORO staining solution (G1260, Solarbio, China) for 20 min. The staining solution was discarded, and the cells were washed 3 times with distilled water. Hematoxylin was added to counterstain the nucleus for 2 min. Finally, ORO buffer was added for 1 min, and the cover glass was sealed with distilled water. For tissue experiments, frozen sections of the spinal cord were dried and washed with PBS, stained with ORO staining solution for 10 min, treated with 60% isopropanol for 5 min, and finally washed with distilled water. Representative images were obtained by fluorescence microscopy.
5
Histological Assessment of Supraspinatus Muscle
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The supraspinatus (n = 5 per group) muscles were flash-frozen and cryosectioned as described previously.29 (link) Masson trichrome (G1345, Solarbio, China) and Oil Red O (ORO; G1260, Solarbio) stains were used to assess fibrosis, atrophy, and fatty infiltration in supraspinatus muscles. Slides were covered with 10% glycerol in PBS (for ORO) or 50% resinene in xylene (for Masson trichrome) and observed on an optical microscope. Cross-sections were chosen randomly from mid-bellies of supraspinatus. Pictures were analyzed with ImageJ software (National Institutes of Health, USA) as described previously.29 (link),30 (link)
6
Histological Analysis of Liver Tissue
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Livers were fixed in 4% paraformaldehyde solution and embedded in paraffin. 5 μm sections were mounted on slides and stained with hematoxylin and eosin (H&E) and Masson, according to the manufacturers’ instructions (G1120 for H&E, G1340 for Masson staining, Solarbio, Beijing, China). For Oil red O staining, frozen sections of the livers were first obtained, and the staining processes were performed according to the manufacturers’ instructions (G1260, Solarbio, Beijing, China).
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