Human epidermal keratinocytes (HEK) are used extensively in our laboratories as an in vitro model of human epidermis to study the vesicant action of sulfur mustard (SM). These adherent cells are released from their tissue culture vessels by proteolytic treatment that detaches the HEK from their plastic supports. However, this common procedure may compromise cells that are struggling to survive and influence the interpretation of damage from these moribund cells. Biomarkers of damage are isolated by lysing cells using specialized buffers or by subjecting the cellular pellet to sonication or other means of disruption. It is especially critical to be able to lyse cells in the desired buffer in situ to minimize losses of biological material used in analysis. Using a Misonix® tissue culture plate sonicator, both 24- and 96-well plates of HEK were subjected to 30-sec bursts in a 4oC tray horn ultrasonic bath at the maximum setting of 10. There was 1 min of cooling between bursts for total sonication times of 2 to 5 minutes. The plates were inspected by microscopy, and wells were photographed to give a rough measurement of HEK lysis. Sonicated plates were also checked for the number of viable cells remaining on the plate or in the supernatant by an MTS-PMS chromogenic viability assay. The supernatant fluid was removed from the well and wells were again checked for the percentage of remaining adherent cells by microscopy. Glutathione levels in the supernatant fluid by both sonicated HEK and acid-extracted HEK were measured and appeared identical. From these results, it appears that the sonication technique may be a useful alternative for the proteolytic method commonly used for the isolation of specific biochemical markers.