Faint bands on PVDF blot.

nsn · #1 (**permalink**) 04-09-2008, 06:58 AM

Hi,

We are constantly getting faint bands despite increasing our primary antibody concentration (1:500 polyclonal Bcl2 ) and loading 50 micro grams of sample into the wells. Do you have any suggestions to resolve this problem, please.
Thanks you !

oBWhat · #2 (**permalink**) 04-09-2008, 02:12 PM

Hello,
this could be several reasons.

1) your protein is limited in concentration (? 50 micrograms should be enough though)
2) your protein is degraded/dephosphorylated
3) your antibody may have been old/degraded
4) you may have inhibiting factor - PBS or something else in your samples

Can you give us more information on how you lysed your sample?

Also have you tried to blot for another protein ie actin or tubulin to check protein?

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Faint bands on PVDF blot.

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