Background in immunostaining. Help.

Giorgio Spagnol spagnol at
Sat Aug 5 10:03:23 EST 1995

Dear fellow scientists,
I'm trying to immunostain Myelin with the serum of a patients having
serum's IgMs toward a myelin-specific glycoprotein.
However, my second, FITC Ab always give me serious problem of background,
even at a 1:3000 dilution. 
here is the protocol.
Fixation in METOH at - 20 degrees for 1'.

1 h at RT in goat serum 1:1 in PBS ( second Ab is goat too )

O.N. at + 4 degrees with first Ab

2 h at RT with second Ab

Any suggestion would be greatly appreciated

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