Injury and inflammation in the gastric epithelium can cause disruption of the pathways that guide the differentiation of cell lineages, which in turn can cause persistent alterations in differentiation patterns, known as metaplasia. Metaplasias that occur in the stomach are associated with increased risk for cancer. Methods for isolating distinct gastric epithelial cell populations would facilitate dissection of the molecular and cellular pathways that guide normal and metaplastic differentiation. Here, we identify Alanyl Aminopeptidase (ANPEP; aka CD13) as a specific, surface marker of zymogenic chief cells (ZCs) in the gastric epithelium. We show that: 1) among gastric epithelial cells, ANPEP expression is confined to mature ZCs, and 2) its expression is lost en route to metaplasia in both mouse and human stomachs. With this new marker coupled with new techniques we introduce for dissociating gastric epithelial cells and overcoming their constitutive autofluorescence, we are able to reliably isolate pure populations of ZCs, and observe changes in ZC differentiation ex vivo in response to epithelial damage.