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In this work, we have focused on 8-methoxypsoralene (8-MOP) complexed with G2.5 and G3.5 poly(amido amine) (PAMAM) dendrimers. The purpose of this study was to investigate the efficacy of half-generation G2.5 and G3.5 PAMAM dendrimers conjugated with 8-MOP for delivery of 8-MOP in vitro study through polivinyldifluoride membrane (PVDE) and prepared pig ear skin (PES) using Franz diffusion and in vivo study through the skin of experimental animals (hairless rat skin). The tissue concentration of 8-MOP in hairless rat skin was analyzed by high performance liquid chromatography (HPLC) after 1 and 2 h. Detailed distribution of 8-MOP in skin layers and cellular structures were analyzed using laser scanning microscopy (CLSM). In vitro and in vivo studies showed that half-generation G2.5 and G3.5 PAMAM dendrimers are able to facilitate transdermal delivery of 8-MOP. G2.5 PAMAM dendrimer appeared to be more effective 8-MOP penetration enhancer than G3.5 PAMAM dendrimer, but in vivo the differences are not statistically significant. The concept of using G2.5 and G3.5 PAMAM dendrimers as carriers seems to be a promising method for the delivery of 8-MOP for PUVA (psoralen-UV-A) therapy.