Freezing down pieces of mammary gland tissue for later transplantation


1. Excise tissue to be frozen taking care to remove any lymph nodes or excess fat pad not containing outgrowth or tumor.

2. Mince tissue into approx. 1-mm-sized pieces (the size that you would eventually transplant).

3. Place tissue in medium containing 7% DMSO and 2% FBS (we normally use RMPI-1640 media since that is what HC11 cells grow in).

4. Slowly freeze aliquots, lowering temperature at the rate of 1ºC/min. Generally, the minced tissue from one inguinal fat pad will yields two aliquots.

5. For short-term storage, freeze to -80ºC and for longer-term storage, store in liquid nitrogen.


1. Quickly melt frozen aliquots by submersing in a 37ºC water bath.

2. Add extra medium to dilute concentration of DMSO. Note that fragments of mammary tissue will not usually pellet by centrifugation because of the high fat content; herefore, fragments of tissue outgrowths and tumors may be collected with forceps.

3. Transplant pieces into the cleared fat pads of 3 week old recipient mice.