Frozen Skin Tissue Block Preparation: A Protocol to Preserve Cutaneous Melanoma Samples from Murine Dorsal Skin

- Quiescent melanocytes stem cells, in response to UV radiation or hair depilation, get activated. Activated melanoma stem cells can lead to cutaneous melanoma - a form of skin cancer. To study melanoma, we prepare frozen skin tissue blocks to retain the native structures of DNA and proteins during the fixing procedure.

Begin by placing a euthanized melanoma mouse on a surgical bed and remove hair from the dorsal region of the skin above the tail. Excise the skin and cover it in a filter paper. Next, submerge the filter paper containing skin tissue in neutral buffered formalin for the desired time. Formalin penetrates the skin tissue and binds to the amino acids, which causes the cross-linking of proteins, therefore leading to tissue fixation.

Remove the skin tissue from paper and cut the tissue to obtain pieces of the desired size. Now, embed the tissue pieces vertically in a cryomold filled with OCT medium and place the cryomolds on dry ice. OCT is a specimen matrix that solidifies on freezing and forms a gelatinous layer around the tissue specimen holding it in place. Use the frozen blocks for further histological assays. In the following protocol, we will prepare frozen skin ti