I have been reading all the e-mail about IF and I also have a few questions.
I use 3%BSA in 1x PBS for our blocking. For 30 min. Our Primary antibodies,
I usually  leave them on overnight at 4 degrees. Then I use either Cy3 or
FITC conjugated Secondary from Jackson Immuno at a 1:250 dilution. I have
some background staining I would like to try to get rid of. Someone stated
that primary antibodies left on to long will cause background. Do other
people find this to be true also?  I was also wondering if people have
better results using BSA IN 1X PBS or Normal Serum from the secondary host
as a block?  Any other suggestions on how to get rid of background would be
great.
Thank You,

Sally Prouty B.S.,HTL
HHMI/University of Iowa
Kevin Campbell Laboratory
400 EMRB
Iowa City, Iowa 52242
Phone-1-319-335-7703
Fax-1-319-335-6957