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Western blot- bands weak - (Feb/28/2013 )

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sdheeru on Sat Mar 2 00:04:21 2013 said:

Possible things I think could affect are:

-The cassette for developing the blot. He has white cassette (from inside) and mine is black.

possible. white would reflect light, black will absorb. try his white cassette.

-the PVDF wetting time.

not likely, 30 seconds in methanol is more than sufficient to activate the membrane (i think 10 seconds is sufficient)

-The side of gel used against the membrane and ultimately upside for the antibody solutions.

not the side of the gel (both are equivalent) but the side of the blot that you are exposing.

have you determined how well the transfer went (stained the blot with ponceau and the gel with coomassie or silver after transfer)?

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