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Next, a fresh medium containing tested compound was added.
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After 24 h incubation, fresh culture medium containing test compounds at various concentrations were added.
After incubated for 24 h, fresh culture medium containing test compounds at various concentrations were added.
After 2 days, the medium was replaced with fresh medium containing test ligands and incubation continued for 5 additional days.
After 24 hr, medium was refreshed and after another 24 hr, the medium was removed and fresh medium containing test compounds (dissolved in ethanol) was added.
Subsequently the medium was changed to 100 μl of medium containing test substances and the cells were incubated from 24 to 72 hours.
After adherence (4 h) the medium was replaced with 1% CS medium containing test compounds in the absence or in the presence of the angiogenic factors VEGF or the NO donor drug NaNP, and incubated for 48 h.
To test for cytotoxicity, 0.1 mL of the maintenance medium containing the tested samples in serial twofold dilutions was added to each of the wells.
Six hours later the culture media was replaced with fresh medium containing the tested compounds at various concentrations.
Test cells (100 μL, 5 × 104 cells/mL) were seeded into a 96-well microplate and incubated about 24 h for cell implantation; then the supernatant was removed, and 100 μL of fresh medium and 100 μL of medium containing a test compound or vehicle control (DMSO) were added.
After preincubation the buffer was changed to a medium containing the test agents, and the islets were incubated for 90 minutes.
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CEO of Professional Science Editing for Scientists @ prosciediting.com