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The scheme in [6] uses a flow rate computation method for routing, which aims to maximize the network throughput.
This methodology employs as mobile phase the same PBS solution used to wash the mallein pre-concentrate in the purification process and uses a flow rate of 0.5 ml/min., an injection volume of 50 μL and a UV detector set to 210 nm.
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Use a flow rate of 1 ml min−1 at 25 °C, and determine the absorbance at 265 nm.
Peptides were separated using an Ascentis Express 150 × 2.1 mm, 2.7 µm C18 column (Sigma-Aldrich, Poole, U.K). using a flow rate of 0.21 mL/min.
We used a flow rate of 1 ml/min with triethylammonium acetate (TEAA)/acetonitrile (ACN) buffer system (solvent A: 100 mM TEAA, pH 7; solvent B: 100% ACN).
Binding analysis was performed by injecting 100 nM of each aflibercept sample over the immobilized surface using a flow rate of 50 µl/min and 10 mM NaOH was used to regenerate the CM5 surface.
A threshold was set on green fluorescence at a value of 200, and samples were analysed using a flow rate below 1,000 events per second to avoid coincidence of viral particles32.
Peptides were eluted using a flow rate of 275 nl min-1, and a ternary gradient, as described62 was used, with the following mobile phases: A (water/acetonitrile/formic acid, 95/5/0.1, v/v/v), B (water/acetonitrile/formic acid, 70/30/0.08, v/v/v), and C (water/acetonitrile/trifluoroethanol/formic acid, 10/80/10/0.08, v/v/v/v).
The volume of the peptide compound injection was 3 μl, using a flow rate of 0.3 ml/min.
Peptides were diluted in 0.1 % formic acid and analysed by direct infusion mass spectrometry using a flow rate of 200 nL/min.
The best separation was obtained when gradient elution was performed and column temperature was kept at 40°C using a flow rate of 0.28 mL/min.
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