Mammary Gland Whole Mount Preparations

 

Day 1: Dissection and Fixation of Mammary Glands

1. Dissect out mammary gland and spread on glass slide.

2. Fix tissue O/N in Carnoy's fixative at RT.

 

Day 2: Rehydration and Staining of Glands

1. Wash in 70% EtOH 2 x 15 min.

2. Wash in 50% EtOH 2 x 10 min.

3. Wash in 30% EtOH 2 x 10 min.

4. Wash in 10% EtOH 2 x 10 min.

5. Wash in distilled H2O 1 x 5 min.

6. Stain O/N in carmine alum stain.

 

Day 3: Clearing and Mounting of Glands

1. Wash in 70% EtOH 2 x 15 min.

2. Wash in 95% EtOH 2 x 15 min.

3. Wash in 100% EtOH 2 x 15 min.

4. Clear in xylene approximately 30 minutes (or until fat is sufficiently cleared from glands).

5. Mount with ~1mL SecureMount and glass cover slip.

 

Carnoy's Fix:

6 parts 100% EtOH

3 parts CHCl3

1 part glacial acetic acid

Carmine Alum Stain:

Place 1g carmine (Sigma C1022) and 2.5g aluminum potassium sulfate (Sigma A7167) in 500mL dH20 and boil 20 min. Adjust final vol. to 500mL with H20. Filter and add thymol crystal as preservative. Refrigerate. Can be used for several months. Discard when color becomes weak.