Objective To investigate the optimal concentration and incubation time of 5-aminole-vulinic acid (ALA) in photodynamic therapy of acne vulgaris. Methods Thirty patients with facial acne vulgaris were equally divided into two groups. In group 1, ALA (10%)cream was applied to acne lesions and protoporphyrin Ⅸ (PpⅨ) fluorescence was examined visually and spectroscopically at 1, 2, 3, 4 and 5 hours. In group 2, ALA cream of 3%, 5% and 10% was applied to lesions in the right cheek, left cheek, and forehead, respectively, of the same patient and incubated for 3 hours followed by photodynamie diagnosis and quantification of fluorescence intensity; clinical outcome and side effects were analyzed. Results Strong brick red PpⅨ fluorescence was observed in inflammatory papules, pustules, and cysts alter applica-tion of 10% ALA cream and irradiation with excitation light. The relative intensity of Ppix fluorescence was 1.3, 4.3, 5.1 and 5.8 in comedones, inflammatory papules, pustules and cysts, respectively, and it increased with the severity of lesions. A higher intensity of PpⅨ fluorescence was noted in patients with longer incu-bation period (3, 4 or 5 hours) compared with those with shorter incubation period (1 or 2 hours), and the difference was significant (P < 0.05). However, there was no significant difference in PpⅨ fluorescence intensity among lesions receiving ALA of different concentrations (P > 0.05). The overall clearance rate was 73% (11/15) after two courses of ALA-photodynamic therapy (PDT) in group 2. Side effects mainly included mild to moderate erythema, swelling and little exudation (occasionally). A transient pigmentation was observed in 2 patients. Neither ulceration nor scarring was noted. Conclusions ALA-PDT is suitable for the manage-ment of acne vulgaris mainly characterized by inflammatory papules, pustules and cysts. The results strongly suggest that 3% and 3 hours are the optimal concentration and incubation time of ALA in PDT of acne vulgaris.