Immunofluorescence stopped working! - (May/02/2011 )

I developed procedure for CK19 and Mammaglobine staining and I had gut results, no background, no unspecific staining in izotyp contorls (almost). After a month I performed the same experiment and know I've got almost no staining in positive control and high staining in only secondary Ab control. (I didn't mix slides!). I used fresh blocking serum (10% FBS) and now my positive staining looked better but secondary ab control gives strong signal (as strong as positive control). Previously I made this experiments many times and it was never like this time!
What can I chack? What could it be?

I would be grateful for any help/advise!

Could your secondary be contaminated with bacteria?

bob1 on Tue May 3 00:06:23 2011 said:

I try to check this. But it looks as usually.
Thank you.