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The following day, these transiently transfected cells were divided into two flasks that were either treated or not with IFN-γ for 48 hrs.
When the OD600 of the cultures reached 0.4 to 0.5, the cultures were divided into two flasks and 1 mM IPTG was added to one of the flasks.
Cultures were grown in PYE medium up to mid-log phase, divided into two flasks, and treated with either 100 μM FeSO4 or 100 μM DP for two hours.
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The solution was mixed for 1 h maintaining a constant temperature of 60°C and finally divided into three flasks.
24 h later the cells were divided into four flasks, each containing 1 × 10 cells for each experiment (in the presence or absence of HAGE).
The cells were centrifuged, washed twice in phosphate-buffered saline, pH 7.0 (PBS: 8 g L−1 NaCl, 0.2 g L−1 KCl, 1.44 g L−1 NandPO4 and 0.24 g L−1 KH2PO4), and divided into nine separate flasks containing 50 mL of mineral medium (MM).
For the formation of noncross-linked and cross-linked PNIPAAm shells on Au NWs, the above Au NWs at initiator solution (8 mL) was equally divided into two separate round-bottomed flasks (labelled with I and II).
This one-way MLR was divided into two 25 cm−2 tissue culture flasks, which were incubated at 45° to the horizontal at 37°C for 5 days.
After nine weeks of incubation, the culture flasks were divided into two groups.
Flasks were divided into two groups to be incubated in the presence and in the absence of cytokines, respectively.
At 8 00 h of the fifth day, the flasks were divided into two groups: (1) cells were subject to a medium change without hormone (control); (2) cells were subject to a medium change containing 10−9 M α-MSH (Sigma-Aldrich, St . Louis MO, USA).
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