The tissue sections and slides were deparaffinized and hydrated. The antigens were heat-retrieved, followed by tissue blocking; overnight primary antibody incubation at 4 °C; secondary antibody incubation; and incubation with ABC reagent. The slides were scanned using the Fluorescence Inversion Microscope System (Olympus, Japan). The tissue sections (5 mm) were stained with hematoxylin and eosin (H&E). The slides were imaged using the Fluorescence Inversion Microscope System (Olympus, Japan).
Fluorescence inversion microscope system
Manufactured by Olympus
Sourced in Japan, United States
The Fluorescence Inversion Microscope System is a laboratory instrument designed for fluorescence imaging and analysis. It utilizes an inverted optical configuration to provide high-resolution visualization of fluorescently labeled samples. The system includes a specialized illumination source, filter sets, and a sensitive camera to capture detailed fluorescence signals from the specimen.
Automatically generated - may contain errors
Lab products found in correlation
Hif 2α, by Abcam (1 mentions)
Digital camera, by Nikon (1 mentions)
Ptch1, by Abcam (1 mentions)
Apollo staining reaction liquid, by RiboBio (1 mentions)
5 ethynyl 2 deoxyuridine edu, by Thermo Fisher Scientific (1 mentions)
Ab41928, by Abcam (1 mentions)
Cd11b antibody, by Bioss Antibodies (1 mentions)
Dapi staining solution, by Beyotime (1 mentions)
14 protocols using fluorescence inversion microscope system
1
Immunohistochemical Analysis of Tissue Sections
2
HIF-2α Immunohistochemistry in Kidney Tissue
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The tissue sections were deparaffinized and rehydrated. For retrieving antigens, slides were heated in 10 mmol/L sodium citrate-hydrochloric acid buffer (Beyotime Biotechnology, China), and then incubated with blocking buffer (10% horse serum and 3% Triton X-100) at room temperature for 2 h. Afterward, the kidney sections were incubated with primary antibody HIF-2α (Abcam, Cambridge, MA, 1:300) at 4°C overnight, then incubated with secondary antibody for 2 h at room temperature. The DAB kit was used to visualize the antibody binding in the kidney sections. The nucleus was stained with haematoxylin as a counter stain. Photomicrographs were taken by Fluorescence Inversion Microscope System (Olympus Corporation, CKX41).
3
Intranasal Delivery of RhB-HupA-NE in Rats
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Animals were anesthetized with diethyl ether, then administered 50 μL of RhB-HupA solution, RhB-HupA-NE, or Lf-RhB-HupA-NE intranasally (Figure 1 ). After 1 h, the animals were anesthetized, and their hearts were perfused with saline. The brains were removed, fixed in 4% paraformaldehyde for 48 h, then placed in a 15% sucrose PBS solution for 24 h until subsidence, then in 30% sucrose for 48 h until subsidence. The brains were frozen at −80 ºC in optimal cutting temperature embedding medium, then cut into 20μm sections using a freezing microtome (Fluorescence Inversion Microscope System, Olympus, Tokyo, Japan).The sections were stained with 300 nM DAPI for 10 min at room temperature, then immediately examined under the fluorescence microscope after washing twice with PBS (pH 7.4). The images were captured using a digital camera (Nikon, Tokyo, Japan).![]()
RhB-HupA encapsulated in NE was intranasally administrated to rats.
4
Mitochondrial Stress Evaluation in EA.hy926 Cells
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EA.hy926 cells cultured on glass coverslips were treated as described above and washed with PBS. After staining with MitoTracker for 30 min and washing with PBS, the cells were stained with DAPI for nuclear counterstaining. The stained slides were photographed using a fluorescence inversion microscope system (Olympus, Tokyo, Japan), the cells were randomly selected to measure mitochondrial injury.
5
Immunofluorescence Analysis of Hedgehog Pathway
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The tumors were fixed with 4% paraformaldehyde and embedded in paraffin. The sections (5μm) were incubated with antibodies: HBx, SHH, PTCH-1, SMO, and GLI-1 (Abcam, USA). The pictures were captured by Fluorescence Inversion microscope system (Olympus, USA)
6
Immunofluorescence Analysis of eIF4E and Ki67
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After the indicated treatment, the cells were collected, washed with PBS twice, and then fixed with 4% paraformaldehyde for 60 min. 1% Triton X-100 in PBS was added to increase the permeability of the cell membrane. After washing with PBS twice, the cells were blocked with 5% w/v BSA/PBS at room temperature for 60 min. They were then incubated with primary antibodies against eIF4E or Ki67 diluted in 5% w/v BSA/PBS (1:200) overnight at 4 °C, and for 60 min at room temperature. Subsequently, the cells were washed twice with PBS and incubated with fluorescent-labeled secondary antibodies diluted in 5% w/v BSA/PBS (1:200) in the dark for 60 min. Finally, after being stained with DAPI for 5 min, cells were observed under a Fluorescence Inversion Microscope System (OLYMPUS, Tokyo, Japan) to detect the fluorescence intensity.
7
Cell Proliferation Assay with EdU Labeling
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Thymidine analog 5-ethynyl-2'-de-oxyuridine (EdU) (Invitrogen; Thermo Fisher Scientific, Inc.) was used to detect cell proliferation. According to the manufacturer's protocol, 2.0x104 cells/well were seeded into 96-well plates and incubated at 37˚C in a 5% CO2 incubator for 24 h. Then, 25 µM EdU was added for 2 h at 37˚C. Following washing with PBS, cells were fixed with 4% para-formaldehyde for 30 min at room temperature and terminated with 2 mg/ml glycine. The Apollo staining reaction liquid (catalog no. C10310; Guangzhou RiboBio Co., Ltd.) was then added to the wells and incubated in dark at room temperature for 30 min. Subsequently, 100 µl 0.5% TritonX-100 was applied to reduce the dye background. DAPI was diluted and used to dye the nucleus in dark at room temperature for 30 min. The results were observed and photographed using a fluorescence inversion microscope system (magnification, x100; Olympus Corporation).
8
Wound Healing Assay in HepG2-HBx Cells
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HepG2-HBx cells could be scratched at interval 0.5cm in 6-well plate and were treated with CP (1/2 IC50; 1/4 IC50, respectively) and cyclopamine (70μM) for 48 h, respectively. HepG2-NC cells were treated without any medicine. The cells could be pictured by Fluorescence Inversion microscope system (OLYMPUS, USA).
9
Immunofluorescence Analysis of PPARγ in 4T1 Cells
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For immunofluorescence analysis, 4T1 cells were washed twice with PBS, fixed in 4% paraformaldehyde, and then incubated with anti-PPARγ monoclonal antibody (ab41928, Abcam, Cambridge, UK) overnight at 4°C. Immunoreactive proteins were detected by incubating with TRITC-conjugated IgG. The nuclei were stained with DAPI (50 μg/mL) for 5 minutes. Images were assessed using a fluorescence inversion microscope system (Olympus, Tokyo, Japan).
10
Histological Analysis of Tumor Tissues
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Tumour tissues were fixed with 4% paraformaldehyde (24 h), and embedded in paraffin blocks for routine histology. Haematoxylin and eosin (H&E) and immunohistochemistry (IHC) staining were performed according to a standard procedure. Human monoclonal LDHB, KI67 and CD11b antibody (Bioss, China) for IHC were used at a 1:300 dilution. The pathological changes were assessed and photographed under Fluorescence Inversion Microscope System (OLYMPUS).
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