Cells in a single-cell suspension were counted with a TC20 automatic counter (Bio-Rad, Hercules, CA, USA). Equal numbers of cells were cultured in triplicate on 24-well plates (CytoOne, USA Scientific, Ocala, FL, USA) for 7 days. Cell proliferation was assessed every 2 days by crystal violet staining, as we described previously [23 (link)].
24 well plate
Manufactured by USA Scientific
Sourced in United States
24-well plates are a type of cell culture vessel used in laboratories. They have 24 individual wells, each capable of holding a small volume of liquid or cell culture. These plates provide a standardized format for various experimental procedures, such as cell growth, drug screening, and biochemical assays, where multiple conditions or replicates can be tested simultaneously.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by Thermo Fisher Scientific (4 mentions)
Dmem f12, by Thermo Fisher Scientific (2 mentions)
Tc20 automatic counter, by Bio-Rad (1 mentions)
Hyclone defined fbs, by GE Healthcare (1 mentions)
Hepes, by GE Healthcare (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
T75 flask, by Corning (1 mentions)
Poly 2 hydroxyethyl methacrylate poly hema, by Merck Group (1 mentions)
Trypsin, by Thermo Fisher Scientific (1 mentions)
Ascorbic acid, by Merck Group (1 mentions)
Dexamethasone (dex), by Merck Group (1 mentions)
Proox model p110, by Biospherix (1 mentions)
5 protocols using 24 well plate
1
Cell Proliferation Assay with Crystal Violet
2
Nanoporous Polymer Membrane Electroporation
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The nanoporous polymer membranes of the LEPDs were first coated with appropriate extracellular matrix protein (fibronectin for cell lines and fibroblasts at a concentration of 1 to 5 μg/cm2; vitronectin for hiPSCs at a concentration of 0.1 to 1 μg/cm2) and incubated for 1 hour. The devices were left uncoated for cells in suspension. Following this, the devices were washed with PBS twice. For adherent cell types, 5000 to 20,000 cells of interest were pipetted into the LEPD wells in 100 μl of the corresponding culture medium. The cells were then cultured on the polymer surface overnight in an incubator (at 37°C with 5% CO2) to promote cell adhesion and tight nanopore–cell membrane contact before electroporation the next day. For suspension cells, 15,000 to 40,000 cells were introduced in the LEPDs containing EP buffer and centrifuged at 150g for 5 min to establish tight cell contact with the nanopores before electroporation. After electroporation, the LEPD arrays were transferred to 24-well plates (USA Scientific) with the appropriate medium depending on the cell type, in an incubator, and cultured for downstream imaging or assays. All experiments were performed on cultures that were passaged less than 10 times.
3
Endometrial Stromal Fibroblast Isolation and Culture
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Stromal cell suspensions from the endometrium were cultured in T-75 flasks (Corning Life Sciences) in stromal defined media consisting of DMEM/F12 (Thermo Fisher) supplemented with Hyclone Defined FBS (GE Life Sciences), Penicillin–Streptomycin (Thermo Fisher), l -Glutamine (Thermo Fisher) and HEPES (GE Life Sciences). Once cells reached confluence, they were passaged using trypsin–EDTA and grown to confluence again. After 3–4 passages a purified population of stromal fibroblasts remained, characterized as vimentin + , CD90 + , and CD45- as previously shown3 (link). Stromal fibroblasts were then plated in either (i) 24-well plates (USA Scientific, Ocala, FL) or (ii) 96-well image-locked plates (Essen Biosciences); and grown for 48 h in stripped media prior to progestin or hormone treatment.
4
BMSC Isolation and Osteogenic Differentiation
5
Hypoxia's Impact on Zebrafish Development
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A hypoxia chamber (ProOx Model P110, BioSpherix, Parish, NY, USA) was used to create a hypoxic laboratory environment in which oxygen was reduced by nitrogen gas (HP, Airgas, Hyattsville, MD, USA). At 30 hpf larvae were exposed to hypoxia (1 mg/L oxygen) for 18 hr or to normoxia (outside of the chamber). Larvae were placed individually in wells of a 24-well plate (CytoOne, USA scientific, Ocala, FL, USA) containing 2 ml clean fish system-water. Twenty-four larvae were used per treatment for surface fish and 48 larvae for cavefish from two separate clutches of eggs per morph. To measure post-anal tail growth, each larva was imaged before placement in a well and again at the end of the treatment using a microscope as described above. Image-J was used to measure post-anal tail length (Figure 6A ) and growth determined as the differential between the two time points. To measure eye size, eye diameter was measured after the 18 hr. hypoxia or normoxia treatment with ImageJ. After imaging, larvae were stained with o-dianisidine and blood cells were counted from images as described above.
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