western blot, I have a signal without primary antibody - (Nov/16/2008 )

Hi I have been using ECL mix to detect the optimum primary Antibody concentrations for a western blot and found out that even without the primary antibody I have a signal for protein blots.
Can anyone shed some light? please!

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Did you use secondary antibody on the blot?

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yes, I did two blots with the primary antibody and without the primary antibody. both blots gave exactly the same intensity signal.

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Seems like your secondary ab is binding to some proteins..

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Are you blocking the membrane properly?

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yes the blocking reagent is part of ECL kit. I block it at RT for 1 hour.

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this must be the answer.

what is your sample anyway? cell lysate?

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