After the tissues were fixed in 10% formalin solution, we embedded them in paraffin blocks and then cut the tissue into paraffin sections. Then, the tissue sections were deparaffinized. Some of the deparaffinized sections were stained with hematoxylin & eosin (H&E), which was used to detect morphologic changes, and some were rehydrated and microwave-heated in sodium citrate buffer (10 mmol/L, pH 6.0) for antigen retrieval. The sections were then incubated with 0.3% hydrogen peroxide/phosphate-buffered saline for 30 mins and blocked with Super Blocking solution (Pierce). Subsequently, the tissue sections were incubated with NCAPG antibody at a 5 μg/mL (ab56382; Abcam, Cambridge, UK) at 4 °C overnight. The sections were washed with phosphate‑buffered saline (PBS) 3 times at 5‑min intervals. Then, they were labeled by EnVision HRP kits (DAKO) at room temperature for 30 mins. Next, the sections were stained with diaminobenzidine (DAB) and hematoxylin dyes and were sealed with neutral resins. All of the sections were observed and photographed with a light microscope with a DP70 CCD system (Olympus Corp.).
Dp70 ccd system
Manufactured by Olympus
Sourced in Japan
The DP70 CCD system is a digital camera designed for microscopy applications. It features a high-resolution CCD sensor that captures detailed images of microscopic samples. The DP70 is capable of capturing images with a resolution up to 12.5 megapixels and can be connected to a computer for image acquisition and analysis.
Automatically generated - may contain errors
Lab products found in correlation
Envision hrp kit, by Agilent Technologies (2 mentions)
Matrigel coated chamber, by BD (2 mentions)
Matrigel matrix, by BD (2 mentions)
Transwell system, by Corning (2 mentions)
Ab56382, by Abcam (1 mentions)
Liquid dab substrate, by Agilent Technologies (1 mentions)
Mayer s hematoxylin, by Agilent Technologies (1 mentions)
Transwell chamber, by Corning (1 mentions)
Matrigel solution, by BD (1 mentions)
8.0 mm pore polycarbonate membranes, by Corning (1 mentions)
Axioskop 2 microscope, by Zeiss (1 mentions)
9 protocols using dp70 ccd system
1
Immunohistochemical Analysis of NCAPG in Tissues
2
Immunohistochemical Detection of UBC9 in Paraffin Sections
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The level of UBC9 in paraffin-embedded tissue sections was detected with immunohistochemical staining. A goat monoclonal antibody was used as the primary antibody. Paraffin-embedded tissues were pretreated at 65°C for 2 h, followed by deparaffinization using standard procedures. Antigen retrieval was carried out with an antigen retrieval solution (10 mmol/L Tris, 1 mmol/L EDTA, Ph 9.0) before 2% sheep serum was added. Then, the slides were incubated with the antibody for UBC9 (ab21193, Abcam) (1:200 dilution) at 4°C overnight. Next, the slides were labeled with EnVision HRP kits (DAKO) at room temperature for 30 minutes, incubated with DAB substrate liquid (DAKO), and counterstained with Mayer› s hematoxylin(DAKO). All of the sections were observed and photographed with a light microscope using a DP70 CCD system (Olympus Corp.).
3
Cell Migration and Invasion Assay
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For the migration assay, 5 × 104 cells were resuspended in serum-free medium and were placed in the upper chambers. For the invasion assays, 1 × 105 cells were seeded in a Matrigel-coated chamber (BD Biosciences, Bedford, MA, USA). After 24 hrs (to examine migration) or 48 hrs (to examine invasion) of incubation, the non-migrated cells on the upper surface of the membrane were removed, and the cells on the lower surface were fixed and stained with 0.1% crystal violet. The cells in 5 random microscopic fields were counted and imaged using a light microscope with a DP70 CCD system (Olympus Corp., Tokyo, Japan).
4
Cell Migration and Invasion Assays
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After seeding for 48h, stably transfected cells were used for in vitro migration and invasion assays. When it came to migration, 6 x 104 cells were plated in the upper chamber with serum-free medium. As for invasion, 1 x 105 cells were placed in a Matrigel-coated chamber (BD Biosciences). After 24 hours (to examine migration) or 48 hours (to examine invasion) of seeding, the upper surface of the membrane was gently wiped to remove unmigrated cells. The cells migrated to the underside were fixed and stained with 0.1% crystal violet. Count cells in five random microscopic fields which use a light microscope with a DP70 CCD system (Olympus Corp., Japan).
5
Transwell Assay for Cell Migration and Invasion
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Migration assays were performed in a 24‐well Transwell chamber with 8.0‐mm pore polycarbonate membranes (Corning, NY, USA). For cell invasion assay, the same membranes in the upper chamber were precoated with 100 μL of 1 mg·mL−1 Matrigel solution (BD Biosciences, San Jose, CA, USA) at 37 °C for 30 min in an incubator. For both migration and invasion assays, 5 × 104 cells in serum‐free medium were placed in the upper Transwell chamber. After 24 h, the cells on the upper surface of the membrane were removed, and the cells on the lower surface were fixed and stained with 0.1% crystal violet. The cells in three random microscopic fields were counted and imaged in a light microscope with a DP70 CCD system (Olympus Corporation, Shanghai, China).
6
Transwell-Based Migration and Invasion Assays
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For the migration and invasion assays, we employed the highly reliable and widely used Transwell chambers. In the migration assay, we introduced 6 × 104 cells into the upper chamber, ensuring the removal of animal serum to accurately assess cell migration. Conversely, in the invasion assay, the upper chamber was precoated with Matrigel, and 1 × 105 cells were seeded to evaluate their invasive potential. After 24 and 48 hours of monitored culture, the cells in the lower chamber were fixed and subjected to staining procedures, specifically tailored for migration and invasion assays, respectively. To capture the results, we employed a microscope DP70 CCD system (Olympus Corp., Tokyo, Japan), capturing random snapshots of cells in 15 fields of view. This approach allowed for a representative sampling and provided a visually compelling representation of the experimental outcomes.
7
Transwell Assay for Cell Migration and Invasion
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Cell migration assay and invasion assays were performed through a transwell system (Corning, NY, USA) with or without Matrigel matrix (BD bioscience) coated above the membrane. Stably transfected cells were suspended in pure DMEM at a concentration of 1x105 /ml. 500μl cell suspension was added in the upper chamber. Fresh medium containing 10% FBS was added in the lower chamber as a chemoattractant. After incubation for 48h, the non-migrated cells on the upper surface of the membrane were removed, and the cells on the lower surface were fixed with methanol and stained by 0.1% crystal violet. The cells in five random microscopic fields were counted and imaged using a light microscope with a DP70 CCD system (Olympus Corp).
8
Immunofluorescence Imaging of Paraffin Samples
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Paraffin-embedded tissue array sections (5μm in thickness) were prepared and immunodetections were detected by immunofluorescence according to the procedures described previously [6 (link)]. The results were visualized and photographed under an Axioskop 2 microscope (Carl Zeiss, Oberkochen, Germany) with a DP70 CCD system (Olympus, Tokyo, Japan).
9
Transwell Assay for Cell Migration and Invasion
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Cell migration assay and invasion assays were performed using a transwell system (Corning, NY, USA) with or without Matrigel matrix (BD bioscience) coated above the membrane. Stably transfected cells were suspended in pure DMEM at a concentration of 1x10 5 /ml. 500μl cell suspension was added in the upper chamber. Fresh medium containing 10% FBS was added in the lower chamber as a chemoattractant. After incubation for 48h, the non-migrated cells on the upper surface of the membrane were removed, and the cells on the lower surface were fixed with methanol and stained by 0.1% crystal violet. The cells in five random microscopic fields were counted and imaged using a light microscope with a DP70 CCD system (Olympus Corp).
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