Abstract: Objective To evaluate the usefulness of protoporphyrin IX （PpIX） fluorescence diagnosis after topical application of 5-aminolevulinic acid （ALA） in human papillomavirus （HPV） related diseases. Methods Photodynamic diagnosis （PDD） was conducted in 36 patients clinically diagnosed as genital condylomata acuminata. PpIX fluorescence was observed 2 hours following the application of 5-aminolevulinic acid 20% cream on the lesions and subclinical lesions of these patients. Biopsy samples were resected from the lesions, subclinical lesions, and normal skin area （0.5 cm and 2 cm around the lesions） of the patients, and subjected to histopathological examination and microarray analysis for HPV DNA. Acetowhitening test was also performed at the four skin areas. Results Of the 36 patients, 30 were diagnosed as condylomata acuminata, 5 as bowenoid papulosis, and 1 as keratosis seborrheica by histopathological examination. Brick-red fluorescence of PpIX was observed in both lesions and subclinical lesions of all patients with condylomata acuminata and those with bowenoid papulosis, in subclinical lesions of 28 patients, at the area 0.5 cm around the lesions of 17 patients, and at the area 2 cm around the lesions of 5 patients. Acetowhitening and HPV DNA were also positive in lesions and subclinical lesions of patients with condylomata acuminata and those with bowenoid papulosis. Mucosa, inflammatory infiltration area and erosion tissue were prone to develop nonspecific PpIX fluorescence. Conclusions ALA-PpIX-mediated PDD can be used for the diagnosis of clinical and subclinical HPV infection, as well as the location of latent HPV infection, however, it′s not recommended to be used in mucosa, inflammatory infiltration area or erosion tissue.