Oil Red O Stain, Isopropanol

(use: Stains neutral lipids)

Product Options:

Part # 1277A 250 ml $50.20 H
Part # 1277B 500 ml $67.20 H

SOLUTION:                                                                                     

250 ml 500 ml
Oil Red O Stain, Isopropanol Part 1277A Part 1277B

 

Additionally Needed:

Alcohol, Isopropyl ACS, 100% Part 12094
Formalin 10%, Phosphate Buffered Part 1090
Hematoxylin Stain, Mayer Modified Part 1202
Lithium Carbonate, Saturated Aqueous Part 12215
Mount-Quick Aqueous Mounting Medium Part 6271A

 

For storage requirements and expiration date refer to individual bottle labels.

 

APPLICATION:

Newcomer Supply Oil Red O Stain, Isopropanol procedure, classified as a physical staining method, is used for identification of fat/lipid in frozen sections.

 

METHOD:

Fixation: Fresh tissue or formalin fixed unprocessed tissue

  1. See Procedure Note #1.

​Technique: Frozen sections cut at 8-10 microns on adhesive slides

Solutions:  All solutions are manufactured by Newcomer Supply, Inc.

All Newcomer Supply stain procedures are designed to be used with Coplin jars filled to 40 ml following the provided staining procedure.

 

PRESTAINING PREPARATION:

  1. Prepare fresh Working Oil Red O Isopropanol Solution; combine and mix well.
    1. Oil Red O Stain, Isopropanol          30 ml
    2. Distilled Water                                 20 ml
    3. Cover solution; allow to stand at room temperature for 10 minutes; filter prior to use.
    4. Solution is stable for 1-2 hours.
  2. Prepare Alcohol, Isopropanol, 60%:
    1. Alcohol, Isopropyl ACS, 100%        60 ml
    2. Distilled Water                                 40 ml
    3. See Procedure Note #2.

 

STAINING PROCEDURE:

  1. Fix frozen section slides in Formalin 10%, Phosphate Buffered for 1 minute.
    1. See Procedure Note #3.
  2. Rinse sections carefully in two changes of distilled water.
  3. Rinse in Alcohol Isopropyl, 60% (Step #2).
  4. Stain in filtered Working Oil Red O Isopropanol Solution (Step #1) for 10 to 15 minutes.
  5. Rinse in fresh Alcohol Isopropyl, 60% (Step #2).
  6. Wash thoroughly in distilled water.
  7. Counterstain with Hematoxylin Stain, Mayer Modified for 2-3 minutes.
  8. Wash gently in several changes of tap water.
  9. Blue in Lithium Carbonate, Saturated Aqueous; 5 to 10 dips.
  10. Wash gently in several changes of tap water.
  11. Blot excess water from slide; coverslip with Mount-Quick Aqueous Mounting Medium.
    1. See Procedure Note #4.

 

RESULTS:

Fat Bright red
Nuclei Blue to dark blue

 

PROCEDURE NOTES:

  1. To freeze formalin fixed unprocessed tissue:
    1. Place specimen in tissue cassette, wash in running water for 5 minutes.
    2. Remove tissue from cassette; blot well, removing all excess water from tissue.
    3. Freeze tissue according to your laboratory protocol.
  2. Isopropyl alcohol must be used; do not substitute another grade of alcohol.
  3. Frozen formalin fixed tissue does not require additional formalin fixation.
  4. Use minimal pressure when applying coverslip or fat/lipid staining may be disturbed.  To remove trapped air bubbles or to recoverslip;
    1. Soak slide in warm water until coverslip is easily removed.
    2. Blot excess water from slide.
    3. Remount with new coverslip and Aqueous Mounting Medium.

 

REFERENCES:

  1. Carson, Freida L., and Christa Hladik. Histotechnology: A Self-Instructional Text. 3rd ed. Chicago, Ill.: American Society of Clinical Pathologists, 2009. 184-186.
  2. Lillie, R. D., and Harold Fullmer. Histopathologic Technic and Practical Histochemistry. 4th ed. New York: McGraw-Hill, 1976. 567.
  3. Sheehan, Dezna C., and Barbara B. Hrapchak. Theory and Practice of Histotechnology. 2nd ed. St. Louis: Mosby, 1980. 205.
  4. Modifications developed by Newcomer Supply Laboratory.