Re:secondary non-specific staining blues

From:Tamara Howard

Hi -

Maybe I've misread your message, but the second post mentioned that a no
secondary antibody control looked positive - that suggests to me that what
you are seeing fluoresce is something in the sample that has nothing to do
with your staining, blocking, washes, what you had for breakfast, etc. Run
a no primary, no secondary (and a no detection if your fluor isn't on the
secondary) and see if that gives your "staining pattern". If so, what you
have do do is kill the autofluorescence...a quick counterstain with
Toluidine Blue might do the trick. I've also seen protocols for osmicating
tissue, UV-treating, or using other dyes (usually blue?) to knowck down

Do you see the same "background" if you do enzymatic detection?

Good luck!


 Tamara Howard
 Department of Cell Biology and Physiology
 University of New Mexico - Health Sciences Center
 Albuquerque, NM 87131

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