western blot sample in high salt concentration - (Jan/23/2010 )

Hi ereryone

My samples are in different salt conentration(0.15M Nacl, 0.5M Nacl, 1.0M Nacl). When i do western to identify them, but i found some bands disappear and some ugly bands. It is Especially strange that the band also disappears in the normal salt concentration.

Is it because the salt concentration affects the result? Should i dilute the sample with water?

Does anyone have apinions on it, please tell me. Thank you very much

salt will have an effect on the running of your samples.

rather than dilute you can dialyze the samples (by drop dialysis) to reduce the salt concentration.

mdfenko on Jan 25 2010, 07:32 AM said:

Thanks, i repeat the experiment and it works.

mdfenko on Mon Jan 25 15:32:02 2010 said:

Can you do drop dialysis on samples already in laemmli buffer?

DNAze on Thu Jul 14 17:56:18 2011 said:

yes. dialyze against laemmli buffer.

mdfenko on Thu Jul 14 18:20:39 2011 said:

I'm doing my own search currently, but do you by chance happen to have a link or a protocol? that would be greatly helpful.

the attached protocol is for nucleic acids but is similar to drop dialysis for proteins. also, you can use standard dialysis membrane instead of filter disks.

in the olden days, we used to dialyze our samples for sds-page against sample buffer (without bromphenol blue or glycerol, which were added after dialysis) prior to loading.