OS cells were transfected with the indicated plasmids. After 48 h, 1 × 103 cells were placed in each well of 96-well microplate for 4 days. Then cells were incubated with 10 µl CCK8 reagent (Liji, Shanghai, China) for 2 h, and absorbance at 450 nm relative to a blank well was measured. For cell migration, transwell and wound healing assays were performed as described previously [17 (link)]. After transfection, 2 × 105 OS cells were cultured in the serum-free medium for 12–16 h and then suspended in serum-free medium. The cells were seeded into the upper chambers and 500 µl complete medium was added into the bottom chambers (Corning, NY, USA) for migration assays. After 24 h, the cells on the lower compartment were fixed in 4% paraformaldehyde (Beyotime) and stained with crystal violet (Solarbio), then photographed and counted with an optical microscope (Olympus, Tokyo, Japan). For the wound healing assay, cells were seeded in 6-well plates at a density of 1 × 106 cells per well. a straight scratch was made using a 200 µl pipette tip when the density reached approximately 100%. After washing with PBS, the loose cells were removed and cells were cultured with serum-free medium. Images were taken with an optical microscope (Olympus) each day for 3 days. Image J software was used to measure the relative wound areas.
Optical microscope
Manufactured by Olympus
Sourced in Japan, United States, China, Germany, Italy
The Olympus optical microscope is a precision instrument designed for detailed observation and analysis of small-scale specimens. It utilizes advanced optical technology to magnify and illuminate samples, enabling users to examine fine details and structures not visible to the naked eye. The core function of this product is to provide a reliable and high-quality platform for microscopic examination across a variety of scientific and research applications.
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Lab products found in correlation
Matrigel, by BD (37 mentions)
Crystal violet, by Merck Group (27 mentions)
Image pro plus 6, by Media Cybernetics (20 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (16 mentions)
Crystal violet, by Beyotime (15 mentions)
Crystal violet, by Solarbio (15 mentions)
Hematoxylin, by Solarbio (12 mentions)
Transwell chamber, by Corning (12 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (10 mentions)
Matrigel, by Merck Group (9 mentions)
Microtome, by Leica camera (8 mentions)
Abc kit, by Vector Laboratories (7 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (7 mentions)
Matrigel, by Corning (7 mentions)
Fluorescence microscope, by Olympus (7 mentions)
Fetal calf serum (fcs), by Merck Group (7 mentions)
3 3 diaminobenzidine (dab), by Merck Group (6 mentions)
Ab15580, by Abcam (6 mentions)
Transwell chamber, by BD (6 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (6 mentions)
1 246 protocols using optical microscope
1
Cell Viability and Migration Assays for Osteosarcoma
2
Evaluating Tumor-Induced Angiogenesis Inhibitors
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A tumor cell-induced chemoinvasion assay was performed using an in vitro co-culture system based on the chemoinvasion assay [48 (link)]. The Hep3B cells were seeded in the lower chamber and treated with 10 and 20 μM of α-mangostin, Man-3DG, and Man-6DG for 24 h. The medium in the lower chamber was replaced with fresh medium, and serum-starved HUVECs (8° × 104 cells/well) were added in the upper chamber. The chamber was incubated at 37 °C for 18 h. The HUVECs that invaded the lower chamber were fixed using 70% methanol and stained with hematoxylin and eosin (H&E) at room temperature for 5 min. The total invaded cells were photographed and counted using an optical microscope (Olympus) at a 100× magnification.
To assess the effects of α-mangostin and its glycosides on tumor cell-induced capillary tube formation, a conditioned medium was collected from the Hep3B cells that were treated with 10 and 20 μM of α-mangostin, Man-3DG, and Man-6DG for 24 h, and used as the angiogenic stimuli for the tube formation of HUVECs. Serum-starved HUVECs (2 × 103 cells/well) were seeded on a surface containing Matrigel (10 mg/mL) from an angiogenesis kit (Ibidi GmbH), and treated with the conditioned medium (CM) for 6 h. Tube formation of the HUVECs was photographed and counted using an optical microscope (Olympus) at a 100× magnification.
To assess the effects of α-mangostin and its glycosides on tumor cell-induced capillary tube formation, a conditioned medium was collected from the Hep3B cells that were treated with 10 and 20 μM of α-mangostin, Man-3DG, and Man-6DG for 24 h, and used as the angiogenic stimuli for the tube formation of HUVECs. Serum-starved HUVECs (2 × 103 cells/well) were seeded on a surface containing Matrigel (10 mg/mL) from an angiogenesis kit (Ibidi GmbH), and treated with the conditioned medium (CM) for 6 h. Tube formation of the HUVECs was photographed and counted using an optical microscope (Olympus) at a 100× magnification.
3
Histopathological Analysis of Spinal Epidural Fibrosis
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The rats from each group were sacrificed and histopathological analysis was performed at week 4 following surgery. The specimens, including the entire L1-L2 spinal column, the paraspinal muscles and epidural fibrotic tissue, were fixed in 4% paraformaldehyde and decalcified, then dehydrated and embedded in paraffin. The gross specimens were then sectioned at a thickness of 5 µm. The slides of each disc were stained with haematoxylin and eosin (H&E; Beyotime Institute of Biotechnology, Inc.) and Masson's trichrome stain (Beyotime Institute of Biotechnology, Inc.) at room temperature. Epidural scar adhesion was evaluated under an optical microscope (Olympus Corporation; ×40 magnification), and the number of fibroblasts was counted under an optical microscope (Olympus Corporation; ×200 magnification).
4
Evaluating SBF-1 Effects on Cell Migration and Proliferation
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A total of 2 × 105 cells per well were seeded into 6-well plates and incubated with DMSO or various concentrations of SBF-1 for 48 h. For migration assay, the cells were washed once with serum-free DMEM and diluted to a concentration of 1 × 105 per ml, and 0.1 ml cell suspension was added to the inner room of transwell (Costar) pretreated with 10 μg/ml fibronectin. Next, 0.6 ml DMEM supplanted with 20% FBS was added into the wells of 24-well plates and the cells were incubated at 37 °C for another 24 h. Then, the cells were fixed with 4% paraformaldehyde (PFA) at room temperature for 30 min and stained with crystal violet (Sigma; 1% in distilled water) for 10 min. After washing two times with distilled water, migrated cells were counted with an optical microscope (Olympus, Shinjuku, Tokyo, Japan; five fields per transwell). For colony formation assay, 1 × 103 cells per well were seeded into 6-well plates and incubated at 37 °C for another 2 weeks. Then, the cells were fixed with 4% paraformaldehyde at room temperature for 30 min and stained with crystal violet for 10 min (1% in distilled water). After washing two times with distilled water, photos were taken with a digital camera (Olympus), and the number of colonies, the cell number of which exceeded 80, were counted with an optical microscope (Olympus).
5
Cell Migration and Invasion Assays
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After transfection, 2 × 105 OS cells were cultured in the serum-free medium for 12–16 h and then suspended in serum-free medium. The cells were seeded into the upper chambers and 500 µL complete medium was added into the bottom chambers (Corning, NY, USA) for migration assays or Matrigel-coated camber for invasion assays. After 24 h, the cells on the lower compartment were fixed in 4% paraformaldehyde (Beyotime) and stained with crystal violet (Solarbio), then photographed and counted with an optical microscope (Olympus, Tokyo, Japan). For the wound healing assay, cells were seeded in 6-well plates at a density of 1 × 106 cells per well. a straight scratch was made using a 200 µl pipette tip when the density reached approximately 100%. After washing with PBS, the loose cells were removed and cells were cultured with serum-free medium. Images were taken with an optical microscope (Olympus) each day for 2 days. ImageJ software was used to measure the relative wound areas.
6
Histological Tissue Analysis Techniques
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Tissue samples were fixed with 10% formalin and dehydrated by various concentrations of alcohol, according to our previous study.34 After embedding in paraffin, tissues were cut into sections of approximately 5 μm. In the process of hematoxylin and eosin (H&E) staining, the slides were incubated in Mayer's hematoxylin and eosin Y solution (Sigma‐Aldrich), and then the images were taken using an optical microscope (Olympus, Tokyo, Japan). For immunohistochemical staining (IHC), the slides were incubated with antibodies against Ki‐67 (Cell Signaling Technology, MA) at 4°C overnight, and then incubated with secondary antibodies at 20‐25°C for 2 hours, followed by mounting with mounting medium (Dako, Glostrup, Denmark). Images were taken using an optical microscope (Olympus). For immunofluorescence analysis, the slides were incubated with antibodies against XIAP or survivin (Abclonal, MA) at 4°C overnight, and then incubated with secondary antibodies Alexa Fluor 488 (Invitrogen), Alexa Fluor 568 (Invitrogen) at 20‐25°C for 2 hours or DAPI (Invitrogen), followed by mounting with fluorescent mounting medium (Dako). Images were captured by using a confocal microscope (Carl Zeiss LSM 780; Jena, Germany).
7
Immunofluorescence Colocalization Analysis
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The 3 mm thick slices were taken and incubated overnight at 4°C with PDGF-B and Cx43 primary antibodies, with a dilution concentration of 1 : 150. Following these primary antibodies incubation, the samples were washed for 5 minutes and then incubated with secondary antibodies at room temperature for 4 hours. Then, an optical microscope (Olympus, Japan) was used to observe the samples, and three ×200 magnifications were randomly selected from each sample. We adopted an immunofluorescence approach to detect the expression of Collagen-I in the atrial tissue. The prepared samples were sealed within 10% serum for 2 hours, then incubated overnight at 4°C with Collagen-I primary antibodies, with a dilution concentration of 1 : 150, and colocalization testing was performed. The samples were then washed and incubated with secondary antibodies at room temperature for 4 hours. DAPI staining was done for 10 min before observation via an optical microscope (Olympus, Japan), with three ×200 magnifications randomly selected from each sample.
8
Thermal Injury Skin Histopathology Protocol
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As previously described [18 (link)], thermally injured skin specimens were fixed in 4% paraformaldehyde (Solarbio, Beijing, China). After the thermal injury, epidermis samples were dehydrated in a series of ethanol gradients and embedded in paraffin, The samples were stained with hematoxylin-eosin (HE) (Beyotime, Beijing, China) and observed under an optical microscope (Olympus, Tokyo, Japan). The number of inflammatory cells infiltrating was identified and counted on every 0.2 mm2 area at regions between epidermis and dermis on HE-stained section with an optical microscope (Olympus, Tokyo, Japan) [19 (link)]. The data were expressed as the mean of all areas counted ± standard deviation of the mean (SEM).
9
Plasma Treatment's Impact on Cell Migration
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A549 cancer cells on 12-well plates were exposed to the plasma for 15 sec in medium. For controls, 2 mm diameter polycarbonate inserts were placed into the 12-well plates, and cells were plated around them. The next day, inserts were carefully removed, and clearance zone was formed. Cellular debris was removed by washing with PBS. Cell migration was documented on days 0, 1, 2, and 3 using an optical microscope (Olympus, PA, USA) with a 4x objective lens (NA 0.10; WD 17 mm). Migration of cells towards the clearance zone (“covering”) was studied by measuring changes in the area of clearance zone using Image J software.
To evaluate the effects of plasma on the surface of 12-well tissue culture plates, media alone (no cells) were placed in 12-grid well plates. Well plates were then treated for 0 (control), 15, and 120 sec, and images of the culture dish bottom surface were taken using optical microscope (Olympus, PA, USA) using a 4x objective lens. After plasma treatment, cells were then added to the well plates and were kept in an incubator for days 1, 2, and 3. Images were taken on each day to observe the cell attachment and growth on the treated surfaces. Gridded 12-well plates were used to properly locate the plasma treated surface. Image J (NIH) software was used to count the cells.
To evaluate the effects of plasma on the surface of 12-well tissue culture plates, media alone (no cells) were placed in 12-grid well plates. Well plates were then treated for 0 (control), 15, and 120 sec, and images of the culture dish bottom surface were taken using optical microscope (Olympus, PA, USA) using a 4x objective lens. After plasma treatment, cells were then added to the well plates and were kept in an incubator for days 1, 2, and 3. Images were taken on each day to observe the cell attachment and growth on the treated surfaces. Gridded 12-well plates were used to properly locate the plasma treated surface. Image J (NIH) software was used to count the cells.
10
Cornea and Conjunctiva Histological Analysis
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Staining (both haematoxylin and eosin and period acid–Schiff [PAS]) was performed as previously described11 (link). Fixed intact eyeballs were dehydrated in an ethanol gradient, cleared in xylene, embedded in paraffin, and sectioned (5 μm thickness) along the sagittal plane. Corneal tissue morphology was observed by haematoxylin and eosin staining and assessed using an optical microscope; conjunctival goblet cell morphology was observed by PAS staining (as described below) and assessed using an optical microscope (Olympus Optical Co. Ltd., Tokyo, Japan). Ten to 15 paraffin sections at matching positions of the anterior segment were stained with a PAS Kit (Sigma-Aldrich, St. Louis, MO, USA) and counterstained with haematoxylin.
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