ARPE-19 cells were seeded at a density of 3 × 105 in 3.5 cm dishes with a silicone insert. After 48 h, the silicone insert was removed, and the cells were exposed to 150 lux blue light and treated with 0.5 µM G570. After the following 24 and 48 h, ARPE-19 cells were observed and recorded by an Olympus IX70 inverted tissue culture microscope (Olympus, Tokyo, Japan) and an SGHD-3.6C charge-coupled device (CCD) imaging system (SAGE Vision, New Taipei City, Taiwan), respectively. The occupied area of ARPE-19 cells was evaluated by ImageJ software (National Institutes of Health, Bethesda, MD, USA).
Ix70 inverted tissue culture microscope
Manufactured by Olympus
Sourced in Japan
The IX70 inverted tissue culture microscope is a high-performance microscope designed for cell and tissue culture applications. It features a sturdy and ergonomic design, and is equipped with advanced optics for clear and detailed observation of samples.
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4 protocols using ix70 inverted tissue culture microscope
1
Blue Light Effects on ARPE-19 Cells
2
Anti-Naegleria fowleri Activity of Bacterial CFS
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In order to confirm the anti-N. fowleri activity of the bacterial CFS and to clearly observe the morphology of the treated amoebae, we prepared 5 × 105N. fowleri trophozoites in T25-cm2 flask containing 5 mL of the complete Nelson’s medium supplemented with 10% FBS, 100 U mL−1 penicillin, and 100 µg mL−1 streptomycin, and incubated at 37 °C for 24 h. Each culture was treated with 1.60 mg mL−1 of the CFS, then incubated at 37 °C for 48 h. LB broth and Nelson’s medium alone were used as the negative controls. Viable trophozoite numbers were determined using the trypan blue exclusion method and the cell proliferation assay, as described above. Total viable amoebae were calculated and shown as mean ± SD and percent of their survival. Morphology of the N. fowleri treated with the bacterial CFSs and controls was observed daily under the Olympus IX70 Inverted Tissue Culture Microscope (Olympus, Tokyo, Japan) and recorded by WiFi Microscope Digital Camera Model MC4KW-G1 (Microscope X, JiangSu, China).
3
Screening Peptides for Anticancer Activity
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Based on the in silico analysis, peptides with antitumor potential were chosen. The peptides were synthesized commercially (GenScript, Piscataway, NJ, USA) and tested for their antitumor activity as described above. HepG2 cells (5 × 103 cells per well) were treated with different concentrations (5.5, 11, 22, 44, 88, 176, and 352 µgmL-1) of each peptide candidate. Cells treated with medium alone and medium supplemented with 1% dimethyl sulfoxide (DMSO) were negative and diluent controls, respectively. The inhibitory activity of each peptide candidate was studied after 24 h of treatment using a CellTiter 96 Non-Radioactive Cell Proliferation Assay (Promega). The inhibitory concentration 50 (IC50) value was calculated using probit analysis. The viable cell number and morphological changes of the treated cells were observed microscopically at 200× magnification using an Olympus IX70 inverted tissue culture microscope (Olympus). Three independent experiments were performed; results are shown as mean ± standard deviation (SD).
4
Anti-tumor Activity of Trichinella spiralis Extract
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The antitumor activity of LE was evaluated in vitro against the three tumor cell lines (HepG2, SK-OV-3, and A549) using the CellTiter 96 Non-Radioactive Cell Proliferation Assay (Promega, Madison, WI, USA). Briefly, aliquots of individual cancer cells (5 000 cells in 100 µL of complete DMEM) were seeded into separate wells of 96-well culture plates (triplicate) and incubated at 37°C in a CO2 incubator for 24 h. The culture fluids were discarded, and the cells were replenished with 100 µL of fresh culture medium containing T. spiralis LE (70 µgmL-1) and incubated for a further 24 h. Medium supplemented with 0.05% (v/v) PIC was used as a negative control. Growth inhibition was evaluated and expressed as percentage cell survival. Three independent experiments were performed, with results presented as mean ± standard deviation (SD). The inhibitory concentration 50 (IC50) value of the LE was calculated using probit analysis with data of percentage cell survival of the tumor cell that was least tolerant to LE treatment (Postelnicu, 2011 ). Morphological changes in the treated cells were observed microscopically (Olympus IX70 Inverted Tissue Culture Microscope, Olympus, Tokyo, Japan) at 100× magnification and recorded by a WiFi Microscope Digital Camera (Model MC4KW-G1, Microscope X, JiangSu, China).
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