Immunohistochemistry-Bad background

PaulKlosen klosen at
Thu Feb 28 10:38:38 EST 2002

oswaldo wrote:

> Have you tried if you get signal without any secondary antibody at all?.
> Sounds like autofluorescence to me...

This is a classical case of autofluorescence, especially if you fix your
tissue with aldehydes. Blood cells are known for this.  The signal should be
there also without any secondary antibodies.

Blocking this is quite difficult, because several techniques to destroy
autofluorescence also end up killing immunoreactivity.

Take a look at two recent papers in J.Histochem.Cytochem ( on
this problem 50(2): 235-244 and 50(3): 437-439.

One solution seems to involve "photobleaching" the autofluorescence on a UV
bench IIRC.

Paul Klosen

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