Autofluorescence

6/2/99

I am a relatively new commer to the world of Confocal Microscopy and

Fluorescence Microscopy and would greatly appreciate your help with a

problem I'm having. Lately I have been trying to perform immunofluorescent

staining on sections cut from a number of differently prepared rat kidney

tissue blocks that we have stored. I have the Fresh Frozen blocks working

perfectly, however the morphology is not very nice. Therefore I was hoping

to use parrafin blocks which have been stored for some time where the tissue

is fixed in paraformaldehyde but not neutralised before embedding. I tried

neutralising the sections after cutting with 150mM Glycine (1 hour at room

temp) but it didn't alter the level of autofluorescence.

Any help with this problem would be fantastic.

Scott Fraser

Immunology Research Centre

St Vincents Hospital Melbourne

FRASERS@SVHM.ORG.AU

There are a few other methods you could try:

1. 50 mM NH4Cl in PBS. Incubate for 15 min at RT.

2. 0.1% sodium borohydride in PBS. Apply while fizzing and incubate 3 x 10

min on ice.

3. 0.15 M ethanolamine, pH 7.5. Incubate 30 min on ice.

Tony Whyte

tony.whyte@BBSRC.AC.UK