Objective: Burn surgeons use autologous skin graft technique for patients; but a challenge remains for large surface wounds. Recently, a method was described which used a small piece of skin to cover a 70 times greater surface by spraying epidermal cells on injured skin. We designed a comparative study to find the best method to make an epidermal cell suspension Materials and Methods: Ten discarded skin samples were sent to our laboratory from Ghotboddin burn hospital, Shiraz. Each sample was sliced into 6 small pieces (1 cm2) and each piece was treated with a different chemical including sodium bromide (2N, 3N, 4N), amunium hydroxide (2N), sodium thiocyanate (2N) and trypsin (0.05%) for 20 minutes. The epidermis and dermis were separated using forceps. Trypsin (2 ml) was added to all samples (except the trypsinized sample) to begin the intercellular detachment. Afterward, epidermis was sliced into small pieces followed by filtration and centrifugation. Identification of keratinocytes and melanocytes was made through immunocytochemical staining for cytokeratin and melanosome antigens, respectively. Results: Incubation of skin with trypsin and sodium bromide resulted in more cell counts and incubation with trypsin had the highest alive cell percentage (p value<0.05). In all methods, some cells were stained positively for cytokeratin antibody and some for melanosome antibody. Conclusion: It is hypothesized that the best technique to make a suspension of epidermal cells is trypsinization since incubation with sodium bromide takes more time and results in a high percentage of dead cells.We are now applying this method on patients.