Analysis of Connexin 43 Expression on Keratinocytes Using Flow Cytometry

Abstract

Single-cell suspensions of primary keratinocytes comprise a heterogeneous cell population that consists of basal cells (stem cells, transient amplifying cells, and post-mitotic basal cells) and suprabasal cells at different stages of differentiation. Quantitative data for the differential expression of epitopes on single cells can be obtained using a flow cytometer. Simultaneous analysis of two intracellular epitopes, keratin 14 and connexin 43, using flow cytometry after keratinocyte isolation, fixation, permeabilization, and fluorescent immunolabeling is described. Three subsets of cells could be distinguished: stem cells (basal cells [keratin 14 positive] that lack connexin 43 expression); suprabasal cells (connexin 43-positive, keratin 14-negative cells); and basal cells (keratin 14 positive) that express connexin 43. The last population of keratinocytes includes both transient amplifying cells and postmitotic basal cells. The scatter characteristics of each cell population are also described.