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The pellet obtained after perchloric acid precipitation was taken up in 300 μl of 0.2 M NaOH and precipitated again by adding an equal volume of 1.2 M perchloric acid.
The AgCl was precipitated again and separated by centrifugation before drying.
The precipitate were redispersed in 10 ml hexane and precipitated again with 20 ml ethanol by centrifugation.
The CNTs were then dispersed in 5 mL of N,N-dimethylformamide using an ultrasonic bath and precipitated again with acetone, filtered, and washed with acetone.
The precipitates were then re-dispersed in hexane and precipitated again in ethanol by centrifugation (3,000 rpm for 5 min).
The precipitate was then dissolved in 10 mL of hexane and precipitated again in 40 mL of absolute ethanol by centrifugation.
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The immunocomplexes were precipitated once again with protein A agarose beads, washed, and eluted in 100 ml of TE with 0.5% SDS and 200 mg/ml proteinase K. Precipitated DNA was further purified with phenol/chloroform extranction and ethanol before amplifying target DNA by reverse transcriptase-polymerase chain reaction (RT-PCR).
After centrifugation, remove 3 mL of the supernatant solution and add 2.5 mL of pH 11.5 NaOH to disperse the precipitates again.
Immunoblotting the precipitates again demonstrated that the P45L mutant cdk2 failed to stably associate with cyclin E whereas this cyclin was readily detectable in the precipitates with the wild type cdk2 construct (Fig. 2C).
To remove excess capping ligands and remaining impurities, the product was again precipitated using ∼5 mL of ethanol and centrifuged at 8000 rpm for 10 min, then redispersed in chloroform.
Afterwards, the supernatant was eliminated and the residual phase again precipitated by 400 µL of hexane.
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