How do I make a blocking buffer solution? - (Mar/25/2008 )

Hi. I am currently having trouble with immunofluorescence staining on cells grown on glass slides (TC-treated). I can do normal primary antibodies with Cy3 secondary, but I have problems with FITC and phalloidin (488). The phalloidin, especially, shows up maybe 20% of the time I use to stain it. I am diluting the antibodies in Dako Antibody Diluent and the phalloidin is diluted into PBS. I'm trying to figure out what is preventing the phalloidin from working properly. Even at 1000ms exposure I still do not see phalloidin.

For blocking buffer, I have access to FBS and 1% BSA solutions - are any of these okay to use?

Thanks,
- Eli

Hello Eli,

In our lab we use 4%BSA and 5%sucrose in Dulbecco's PBS. Works fantastic on polystyrene plates. Our group also manufactures slide arrays. We use 1%BSA and 5%Sucrose in Dulbecco's PBS for the slides. Hope this information helps.

Have a good day.