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Then, a sodium hydroxide solution (6 molar) was added dropwise into the above mixture with vigorous stirring until the pH value reaches 10.
Furthermore, when higher DMAB molar was added to the solution (with orange color only), a new intense absorption band appeared at 410 nm which was indicative of the formation of nanoparticles with spherical shape.
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In Figure 9 (left), it is possible to appreciate the evolution of the maximum absorption band (UV vis spectroscopy) when variable DMAB molar is added to the solution.
The crude peptides were dissolved in AcOH-H2O 4:1 to a final concentration of 2 mg/mL, and iodine (10-fold molar excess) was added in one portion.
The sol stabilizer ethanolamine in 1 1 molar ratio was added in this solution and stirred for the next 1 h at 60 C.
N-Bromosuccinimide (NBS, Sigma) was resuspended in the same buffer and an 80-fold molar excess was added to the LcrV solution.
The antibody or competitor oligonucleotide (the single stranded oligonucleotide in 20-fold molar excess) was added at the start of the incubation for the supershift and competition experiments.
When NTD125 (1∶5 molar ratio) was added prior to denaturation, there was no loss of wild type (PAb1620) (Fig. 3a, panel-2, lanes 2, 5, 8, 11, & 14, see arrow).
The reaction mixture was cooled to 0°C using ice bath and varying amounts of acryloyl chloride (1-1.5 molar excess) was added drop wise using an addition funnel.
For competition experiments, one μl of non-radiolabelled Egr-1 oligonucleotides (20-fold molar excess) was added to the binding mixture before addition of the radiolabelled probe.
UDP-GlcUA in molar excess was added, and the samples were incubated at 37 °C for 60 min. Final reaction concentrations were as follows: 100 μM Tris HCl (pH 7.4 /5 mM MgCl2/5 mM saccharolactone/2% DMSO/250 μM substrate/2 mM UDPGA/50 or 5 μg of total protein, respectively.
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