I'm sure it is associated with Picro-Sirius red as it doesnt occur until I apply this. I originally thought it may be a drying artefact however I have tried not drying the slides and it still occurs. I have also tried leaving the slides longer in alcohol but this didnt help, and if I leave them too long all of the yellow staining is removed from the tissue sections. Even if I do not apply weigerts haematoxylin the artefact can still be seen within some cell nuclei. The actual connective tissue components of the tissue section look fantastic but this can be seen in other areas. It seems to be worse on columnar epithelial cells and liver hepatocytes.
Method is as follows:
0.5%acidified potassium permanganate - 5 mins
1%oxalic acid -1 min
Rinse in alcohol put into Millers solution in 37 degree Celsius oven - 2 hrs
Rinse in alcohol then distilled water
Weigerts hx - 15 mins
0.5% acid alcohol - 3to 5 seconds
Blue in tap water - 3 to 5 mins
Picro-Sirius red - 1 hrs ( I have tried different timings for this stage from 20 mins to 1 hrs and the same problem is encountered. Solution is ready to use not made in house)
Blot dry
DCM ( rapidly through alcohols)
Are you referring to the pigment granules in the hepatocytes in the first two images? If so, is it also present in the H&E sections as faint golden-brown granules? If it is, it could be lipofuscin pigment.
I think that these are crystals of picric acid that can be dissolved in ethanol (when rinsed after fixation with picric acid, for example, in Bouins fluid, 70% is indicated), since it is poorly soluble in water. But if picric acid turns yellow some important areas of the histological section, leave it as it is.
Filter all solutions immediately before applying on slides and try again
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