Mesenchymal stromal cells (MSC) have been introduced into the field of tissue-engineered airway transplantation. Since patients with extensive tracheal defects often require an open tracheotomy, this study investigated if MSC could be obtained from the adipose tissue of the neck during this procedure. Cells were isolated by plastic adherence from the adipose tissue of 8 patients. Cell isolates were analyzed for (i) proliferation, (ii) the expression of CD marker molecules and (iii) multilineage differentiation. The isolated spindle-shaped cells showed a high proliferation capacity and the flow cytometric analysis revealed a distinct population meeting the criteria for MSC. Using classical MSC cultivation protocols the characterized cells showed adipogenic, chondrogenic and osteogenic differentiation for all analyzed cell isolates. This study was able to demonstrate that sufficient amounts of stem/progenitor cells can be easily isolated from adipose tissue of the neck obtained during open tracheotomy. These cells may be a source for future tracheal replacement therapies.