The calcium-activated potassium channel, KCa3.1 is implicated in the process of exocytic release of intracellular granules from certain secretory cells such as human lung mast cells. MUC5AC is a predominant human respiratory mucin that is upregulated in asthmatic human airways, and is hypothesised to be released from specialised secretory human airway epithelial cells, such as goblet cells, via exocytosis of preformed intracellular mucin granules. Whether KCa3.1 channel activity plays a role in mucin granule exocytosis, in a similar to manner as its role in human lung mast cell degranulation, is unknown. The aim of the study was to investigate whether the KCa3.1 channel plays a role in MUC5AC secretion from airway epithelial cells. Expression of MUC5AC and the KCa3.1 channel was investigated in primary human bronchial epithelial cell (HBEC) cultures isolated from bronchoscopy brushings using quantitative reverse transcriptase (RT)-PCR and in bronchial biopsies using immunohistochemistry. KCa3.1 channel activity was examined using patch clamp electrophysiology. KCa3.1 channel mRNA expression was similar in submerged asthmatic and healthy HBECs. Analysis of expression levels in differentiated cultures is yet to be completed. MUC5AC mRNA was detected in differentiated cultures but not in submerged cultures. MUC5AC and KCa3.1 channel immunoreactivity were demonstrated in paraffin-embedded air-liquid interface (ALI) cultures, and in the bronchial epithelium severe, moderate and mild asthmatics, and in healthy controls. Analysis of sequential sections of bronchial epithelium revealed co-localisation of MUC5AC and the KCa3.1 channel in approximately 50% of cells. Analysis of KCa3.1 channel and MUC5AC immunostaining in bronchial epithelium indicated that severe asthmatics exhibited significantly higher levels of KCa3.1 channel (P = 0.0012) and MUC5AC (P = 0.0180) staining. Patch clamp experiments revealed that submerged asthmatic HBECs exhibited significantly larger KCa3.1 currents than submerged healthy HBECs (P = 0.0225). To conclude, the presence of MUC5AC mRNA in differentiated cultures but not submerged cultures indicates that differentiated cultures are the more suitable model of airway epithelial cells for this study. Co-localisation of MUC5AC with KCa3.1 channel protein suggests that mucus-secreting airway epithelial cells express the KCa3.1 channel. Preliminary immunostaining results and patch clamp electrophysiology experiments implicate a possible difference in KCa3.1 channel expression and activity in airway epithelial cells in asthma and in health.