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Smeared Bands after Ponceau Staining?


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3 replies to this topic

#1 Elizabeth Rendina

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Posted 13 July 2013 - 07:56 AM

Hello,

I have observed smeared looking bands on my Ponceau-stained PVDF after transfer. My samples are from bone, so I'm not sure what is the cause for this and/or if I should be concearned. Other than ~3-4 defined bands, the rest just looks smeared... (please see photo)?

I'm not sure if this is what protein degradation looks like, but I do add appropriate protease/ phosphatase inhibitors to my buffer?

Any ideas/ suggestions would be greatly appreciated!

Thanks all,

Beth

Ponceau Bone_1.JPG
Ponceau Bone_1.JPG
Ponceau Bone_1.JPG

#2 bob1

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Posted 15 July 2013 - 01:21 AM

It is quite likely to just be a level of protein thing, you could test by coomassie staining a gel and see if you get the same pattern.

#3 mdfenko

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Posted 15 July 2013 - 03:56 AM

it could also be due to degraded and/or aggregated proteins.

do you boil the samples in the sds loading buffer? if so, try incubating at 60-70C for 10-20 minutes instead of boiling.

Edited by mdfenko, 15 July 2013 - 03:56 AM.

talent does what it can
genius does what it must
i do what i get paid to do

#4 Elizabeth Rendina

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Posted 15 July 2013 - 06:14 AM

Thank You I'll try this today!




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