Isolation of tdTomato Expressing Inter-follicular Epidermal Melanocytes or Keratinocytes from Mouse Tail Skin.

Abstract

The epidermis is the outermost layer of the skin. It is made up of mostly keratinocytes along with a small number of melanocytes and Langerhans cells. Melanocytes produce a pigment called melanin, which is transferred to the keratinocytes, and protects these cells from damage from UV radiation, as well as generating hair and skin colours. In this important relationship, keratinocytes exert control over melanocytes. Many questions regarding keratinocyte-melanocyte interactions have yet to be answered, and would benefit from study in model systems, to address diseases such as vitiligo and cutaneous melanoma. Most of the mouse is covered in fur and these areas lack the skin pigmenting inter-follicular epidermal (IFE) melanocytes. However, the mouse tail is pigmented analogously to human skin. Here, we present a method for isolating IFE melanocytes or keratinocytes expressing the tdTomato marker from the mouse tail, using fluorescence-activated cell sorting (FACS). The method involves firstly separating the tail skin epidermis from the dermis, and then digesting the epidermis to produce dissociated cells, which can then be sorted. These isolated cell populations can be studied using RNAseq or cultured in vitro. This protocol isolates IFE melanocytes or keratinocytes and immediately provides reasonable yields of cells, without the need to stain the cells for cell specific markers.