Healthcare-associated infections (HAI) are illnesses acquired during healthcare and are often the most important adverse event during healthcare. With the aim of increasing the effectiveness of disinfection/decontamination processes in the health service with safe and not promote microbial resistance, we propose the development of portable equipment associated with type C ultraviolet light (UVC). The efficiency of the irradiance emitted by the equipment (at dosages 3.5, 5.0, and 60 mJ/cm2) was determined by the action exerted after exposure against four different bacterial (Acinetobacter baumannii, Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus) and three different fungi (Candida albicans, C. parapsilosis, and Aspergillus section Fumigati). It was possible to observe that all treatments were capable of inactivating the bacterial species evaluated (p < 0.05), causing the irreversible death of these microorganisms. The most effective elimination of fungal agents was at a dose of 60 mJ/cm2 of UVC radiation, with a decrease in the fungal inoculum varying between 94% and 100% in relation to the control without exposure. Thus, our study showed that the application of the portable prototype with UVC light (254 nm) at a distance of 48 mm, allowed an average irradiance of 3.5 mW/cm2, with doses of 3.5 ≈ 60 mJ/cm2 (from 1 to 60 s of exposure), which can promote the total reduction of the bacteria evaluated and significantly reduce fungal growth. Therefore, this prototype could be used safely and effectively in the hospital environment, considerably reducing contamination and contributing to the reduction of healthcare-associated infection risk.