[Histonet] Oil Red O Churukian Method

[Anna K. Schultz]

Hi,
 I am looking for a better Oil Red O protocol, and I found Gayle Callis' 
posting on the Churukian method--but it leaves out the amount of 
isopropyl alcohol in which to dissolve the .5 gm Oil Red O.  I was 
wondering if anyone could tell me this amount.

thanks a lot,
Anna

here is a copy of the post of the protocol from the archives:
Oil Red O/Dextrin, Churukian method
Fresh tissue (not prefixed) frozen sections are immersed immediately 
into Neutral buffered formalin for a minimum of 10 min (can be 
hours/days). Rinse sections with distilled water before staining. 
Prefixed, with NBF should be cryoprotected in 20 - 30% sucrose, mounted 
on Plus Charge slides and air dried for 30 min to 1 hour, or longer to 
insure sections stay on slide. Do not fix frozen sections with alcohol 
or acetone to prevent lipid removal.

Protocol:
1. Immerse dry slides directly into filtered 0.5% Oil Red O in Dextrin , 
stain 20 minutes 2. Rinse VERY GENTLY in running tap water 3. 
Counterstain with Gill II hematoxylin for 20 - 30 seconds 4. Rinse 
gently with water, blue in Scotts tap water type bluing solution, (NOT 
AMMONIA WATER), rinse gently, and coverslip with aqueous mounting media.

Reagents: Dissolve 0.5 gm Oil Red O in absolute isopropyl alcohol and 
stir overnight. Dissolve 1 gm dextrin (bacteriological grade or TYPE III 
(Sigma) from corn in 100 ml distilled water Working solution is 60 mls 
stock Oil Red O and 40 ml 1% dextrin solution Stable for months, and 
reported to work on paraffin sections. Reference: Gamble and Bancroft, 
Theory and Practice of Histological Techniques, 5th Edition, 2001 with 
photos. This method also published in J of Histotechnology by Charles 
Churukian. Go to JOH archives for journal year/volume.

-- 
Anna K. Schultz
SDRC Core A Research Technician
Department of Dermatology
College of Physicians and Surgeons
Columbia University
Vanderbilt Clinic 15-211
630 West 168^th Street
New York, NY 10032
Phone: (212) 305-4954