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problems with SDS-PAGE

SDS-PAGE

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#1 emmahoshi

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Posted 17 June 2014 - 10:34 PM

hello 

 

i've been making SDs-page gels both 12% and 16% for a while now but i had a problem after staining the gel with either silver of coomassie stain which is that a line appears at the lower end of the gel and the problem is that some important bands appear around that mwt and can't be seen because of this random line i use a similar recipe to this one http://mcardle.oncol...preparation.pdf   for the gel so can anyone help me in solving this problem 

 

 

thank you 



#2 mdfenko

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Posted 23 June 2014 - 03:55 AM

the line is probably a "buffer" line. sometimes you can run the gel longer and it will pass the protein (not always) it could also be due to decomposing (read "old") sds.

 

is the line new to you?

 

what is the apparent mw?

 

what is the mw of your protein of interest?

 

you may be able to clear up your problem by switching to tris-tricine sds-page (used for low molecular weight proteins and peptides).

 

post a picture of your gel for a possibly better diagnosis.


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#3 emmahoshi

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Posted 23 June 2014 - 06:54 AM

thanks for the replay my dr. normally uses ready made gels but we found out that clearer bands are with those gels you make your self the line only appears with those gels we make ourselves so yeah the line is somewhat new the problem is that we are purifying a protein that starts at 55kd and stage by stage reaches-8 kd  i'll post the gel pic as soon as i reach the lab thank you 







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