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I’ve prepared slides for immunohistochemistery finished staining with DAP and Heamatoxyline and put the covers lip but unfortunately the staining was poor
I want re-stain the slides is there any protocols for this ???? I want to remove the cover slip (this is my problem) !!!!!!!
thanks
It depends on which mounting medium you have used. For synthetic Canada balsam for instance, putting the slides into a Coplin jar with xylene and some patience will do the trick without physically damaging the sections. For water-soluble media a buffer solution may prove useful. Unfortunately there is no guarantee that your new immunolabeling would work. Repeat all the immunodetection steps as your antibodies and enzyme have surely been damaged. For hematoxylin there should be no problem.
Let us know whether you succeeded.
Best of luck,
D
Removing the coverslip is the hardest part. I soak slides in 60C xylene for 5-10 minutes to dissolve the mounting medium (I use prolong antifade). This loosens the grip of the coverslip. The tissue I use is paraffin embedded, but by the re-staining phase they are already de-waxed. Still I'm not sure if tissue embedded differently will be affected by xylene. (Xylene is a hydrocarbon, in this case used to solubulize hydrophobic mediums, like wax). If the coverslip is still tightly attached, I gently lift a corner with a pipette tip, and squirt some xylene underneath the slip to further loosen it. Then with fingers or tweezers pull it up. If it's not loose enough the tissue might get pulled up with the slip.
I've successfully re-stained IF slides with a second round of IF, or followed by periodic acid Schiff/Alcian blue stained. You can also follow FISH stains with PAS/AB, just not the other way around. I'm not sure how successful DAB/heamotoxyline re-staining would be, I've never tried it.
Hope that helps
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