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EZ-BindShut II

Manufactured by AGC Techno Glass
Sourced in Japan

The EZ-BindShut II is a laboratory equipment product. It serves the core function of binding and sealing sample containers or vials.

Automatically generated - may contain errors

4 protocols using EZ-BindShut II

Cells cultured under monolayer conditions were detached with trypsin-EDTA, and filtered through a 40 μm cell strainer. The cells were then inoculated into 1% methylcellulose-containing serum-free αMEM supplemented with B27 (Life Technologies, Carlsbad, USA), 10 ng/ml human EGF (PeproTech, Rocky Hill, USA) and 10 ng/ml human bFGF (PeproTech), at a density of 5 × 103 cells per well on 6-well-type ultra-low attachment plate (EZ-BindShut II, AGC Techno Glass, Shizuoka, Japan). After 10–14 days incubation, spheres were observed with the assistance of an inverted phase contrast microscopy, and analyzed by BZ analysis software on a BZ-9000 fluorescence microscope (Keyence, Osaka, Japan) using the Hybrid Cell Counting module. Sphere-forming units were determined at a given day by counting cell aggregates with larger than 3,000 μm2 surface area and with the ratio of the longest diameter and the shortest diameter (L/S ratio) less than 1.5 (spherical figure). The whole area in a dish was scanned by automated microscope, and sphere number per dish was calculated from tiled image data.
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MIA PaCa-2 and PANC-1 cells were pre-treated with 200 or 400 μg/mL TiOxNPs for one hour followed by 2 or 5 Gy radiation treatment. Next, cells were trypsinized, counted, and plated in ultra-low attachment 96-wells plates (EZ-BindShut II, AGC Techno Glass, Shizuoka, Japan) at a density of 1000 cells/well. The cells were maintained in serum-free alpha MEM supplemented with B27 (Life Technologies), 10 ng/mL rhEGF (PeproTech), and 10 ng/mL rhbFGF (PeproTech), and then mixed with 1% methylcellulose. The number of spheres over 20 μM was evaluated after 10–12 days using BZ analysis software on a Biorevo BZ-9000 microscope (Keyence, Osaka, Japan).
For the second passage, spheres were dissociated with 500 μL accutase for 5 to 10 days at 37 °C until a single cell suspension was obtained, followed by incubation with 200 or 400 μg/mL TiOxNPs for one hour followed by 2 or 5 Gy radiation treatment. Cells were then cultured and analyzed after 10–12 days.
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Cells in monolayer culture were dissociated, filtered through 40‐µm cell strainer to obtain single‐cell suspension, and seeded at a density of 2500 cells/well onto a 96‐well ultralow adherence plate (EZ‐BindShut II; AGC Techno Glass) containing 1% methylcellulose containing αMEM medium supplemented with B27 (Life Technologies), 20 ng/mL rh EGF (Wako), and 20 ng/mL rh bFGF (Wako). After 14 days of incubation, mammospheres were visualized under an inverted phase‐contrast microscope, and BZ software was used to analyze on BZ‐9000 (Keyence) equipped with a hybrid cell counting module. The experiment was repeated using MMECs obtained from different mouse pairs including littermates.
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Cells cultured under 2D conditions (monolayer) were dissociated with trypsin-EDTA, filtered through a 40 μm cell strainer, and then inoculated into 1% methylcellulosecontaining αMEM supplemented with B27 (Life Technologies), 10 ng/ml rh EGF (PeproTech) and 10 ng/ml rh bFGF (PeproTech), without serum, at a density of 5 x 10 4 cells on 6-well-type ultra-low attachment plate (EZ-BindShut II, AGC Techno Glass). After 14 days incubation, spheres were observed under the inverted phase contrast microscopy, and analyzed by BZ analysis software on BZ-9000 (Keyence) using hybrid cell counting module.
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