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These cells, designated passage 1, were cultivated in growth medium (DMEM+10% FBS+antibiotics) at 37°C in 95% air, 5%CO2.
To generate a working cell bank, primary fetal RPE stocks were expanded approximately ten-fold and these working stocks were designated Passage 0 (P0).
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Cells below passage 10 were designated "early passage cells".
These cells, designated as passage 1 (p1) of SHhES1-NPCs, formed typical neural progenitor rosette structures.
When the attached "primary MEFs", which were designated as passage number 0, reached confluency, they were subcultured every 3 days at a density of 20,000 cells/cm.
The remainder of the primary quadrant was harvested with a sterile cotton swab, resuspended in LBG broth, mixed with an equal volume of 40% glycerol, designated "laboratory passage #1", and stored at -70°C.
Cells isolated from tissue samples were incubated in DMEM/F12 media containing 10% FBS for 10 14 days to allow attachment and the formation of colonies; these primary cultures were designated as passage 0. All cultures were kept in a humidified incubator with 5% CO2 in air at 37°C.
Low-passage LNCaP cells (LNCaP) were designated as under passage 25, whereas high-passage cells (HP-LNCaP) were over passage 75.
For the studies in this report, ATCC 10-87 VERO cells at p148 and p256 were designated as low passage (10-87 LP) and high passage (10-87 HP) VERO cells, respectively.
The virus was isolated in embryonated chicken eggs (ECE) at Biopharma Laboratory in Rabbat, Morocco, followed by passage in BSR cells, a clone of BHK-21 cells (designated isolate 2-451- 2-451-e 1E3BSR).
We designated the cell passages 5, 8, 12 and 22 as 1, 2, 3 and 4 respectively.
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