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Lab Standard Operating Procedure (SOP) for Immunohistochemical Staining
for Filipin
Materials
Literature
 1.
Dissolve 5mg of filipin (Filipin complex from Streptomyces fillipinesis
Sigma-Aldrich product #46329) in 1ml dimethylformamide and add 100ml
PBS buffer.
2. Stain frozen sections (ours were previously fixed and then frozen)
for 30 minutes in the solution. Use frozen adrenal as control.
3. Wash with distilled water and counterstain with hematoxylin for
1 min.then wash in distilled water
4. Do not use acid alcohol, ammonia water is okay
5. Wash in distilled water and
6. Mount with water based mounting media
7. Filipin stains free cholesterol.
8. Filipin-stained sections are digitally photographed using a x40
oil-immersion objective (N.A. 1.35) with 340-nm excitation and a
510-nm emission barrier filter. Images were acquired by use of Axon
Imaging Workbench 2.2 with constant excitation intensity and exposure
(4 s). Mean pixel intensity of three filipin-stained sections was
determined from three control and three hypercholesterolemia animals
(n = 18 total sections) by use of Image J software.
Last
GLP modification 8/15/04:
Douglas
Bowles
Jennifer Casati
James R. Turk
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