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After ingestion of a 75 g glucose load, blood samples will be obtained at 0, 30, 60, 90 and 120 min for determining the glucose and insulin levels.
At time zero, the HBSS solution of the upper compartment was removed and replaced by 0.5 mL the same buffer containing either 50 μg/mL DOX or Asp-DOX; 100 μL solution in the lower compartment was sampled after 60 min for determining concentration.
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The cells were lysed with reporter lyses buffer (Promega) and after centrifugation of 2 min, supernatant was used for determining β-galactosidase normalized luciferase activity by adding 100 μl luciferyl-CoA (Promega) to 20 μl of cell extract in a monolight luminometer (BD Biosciences).
After settling at room temperature (25 °C) for 30 min and then centrifugation at 6000 rpm for 5 min, the supernatant was obtained for determining the glucose solubility by using the dinitrosalicylic acid (DNS) method (Miller 1959).
After centrifugation at 3,000 g for 10 min, the supernatants were collected for determining amounts of pentoses and hexoses released from enzymatic hydrolysis.
The min-max criterion is useful for determining order of the insertion.
The resultant precipitation was removed by centrifugation at 12,000 rpm for 10 min. The supernatant was used for determining the yield of SAM by the HPLC.
Three optimization models, sensitivity-based, scenario-based and min max, are proposed for determining robust optimal improvement schemes that make system performance insensitive to realizations of uncertain demands or allow the system to perform better against the worst-case demand scenario.
Shake the plate in the reader at 306 r.p.m. for 2 min. Incubate for 15 min. Determine the absorbance (493 nm) and the fluorescence (excitation 485 nm; emission 520 nm) at 25 °C.
The reaction mixture was incubated at respective temperatures for 30 min before determining protease activity according to the standard assay procedure earlier described.
The reaction was initiated by the addition of 120 μL of 5 mM ferrozine into the mixture, which was then left at room temperature for 10 min before determining the absorbance of the mixture at 562 nm.
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