|Mammary gland Whole Mounts
Remove inguinal mammary gland and place on a microscope slide. Use blunt tweezers to manipulate the gland, spreading it out so it resembles its orientation in the boys. Immediately place slides in Kahle’s fix.
Allow the glands to fix for a minimum of 4 hours (overnights is fine).
Wash in 70% EtOH for 15min
Gradually change to distilled water
Rinse in distilled water for 5 min.
Stain in carmine alum o/n
Wash in 70% EtOH 15min
Wash in 95% EtOH 15min
Wash in 100% EtOH 15min
Clear glands in xylene and mount with permount under a coverslip
The time required for clearing varies from gland to gland. Some of the larger, fattier glands will take several days, but average adult mammary glands usually take anywhere from a few hours to 1 day. You’ll have to visually judge when they are done clearning. The fat has an opaque appearance and when clearing is complete, the former fat pad should be almost transparent.
Kahle’s fix (500ml)- stored at RT
53 ml 37% formaldehyde
147 ml 95% EtOH
9.8 ml glacial acetic acid
290.2 ml dH2O
Carmine Alum stain
Place 1g carmine (Sigma C1022) and 2.5g aluminum potassium sulfate (Sigma A7167) in 500ml dH2O
Boil for 20min
Adjust volume to 500ml
Add a crystal of thymol as a preservative
Store @ 4˚C
Solution can be reused several times. Discard when color becomes weak.