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Spores were prepared as described for the ISR assays but were finally resuspended in sterile distilled water to the desired final concentration of 106 spores/ml.
Samples spanning 0.1 1.5 OD at A230, A250, or A280 (Supporting Information, SI-Table 3) were prepared in a total volume of 500 μL by mixing concentrated protein solution with reference buffer to the desired final concentration for each experiment.
After complete evaporation of acetone from the Erlenmeyer flasks, 100 mL sterile culture media was added under laminar flow hood so as to reach the desired final concentration of organic compound.
Bufalin entity, blank liposomes, blank PEGylated liposomes, bufalin-loaded liposomes, and bufalin-loaded PEGylated liposomes were diluted into complete medium and added to cells in a total volume of 0.1 mL to achieve the desired final concentration; for the control, no test solution was added.
Drugs were from Tocris-Cookson (Bristol, UK) and Sigma, and were dissolved in the perfusing ACSF at the desired final concentration.
The LUV suspensions were diluted to desired final concentration and then filtered using a syringe filter with 0.45 µm pore size (Whatman, Florham Park, NJ).
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This constant-volume diafiltration step is preceded and followed by optional ultrafiltration or pure dilution steps that force the concentrations at first to arrive to the optimal macro-solute concentration and at last to arrive to the desired final concentrations.
This constant-volume diafiltration step is preceded and followed by ultrafiltration or pure dilution steps that force the concentrations at first to arrive to the optimal macro-solute concentration and at last to arrive to the desired final concentrations.
Varying doses of DOC (with equal total amounts of dimethyl sulfoxide (DMSO), the solvent control) were made up in fresh media and 0.5 mL was added to cells already plated to achieve the desired final concentrations of DOC.
Different concentrations of test samples were obtained from stock solution and placed in separate vials, and the volume of each vial was made up to 5 mL with brine to obtain the desired final concentrations (10, 20, 40, 60 and 80 µg/mL).
Samples of the stock solutions were added to normal saline solution or internal solution to give the desired final concentrations.
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