2469-7311 (c) 2016 IEEE. Personal use is permitted, but republication/redistribution requires IEEE permission. See http://www.ieee.org/publications_standards/publications/rights/index.html for more information. This article has been accepted for publication in a future issue of this journal, but has not been fully edited. Content may change prior to final publication. Citation information: DOI 10.1109/TRPMS.2017.2679010, IEEE Transactions on Radiation and Plasma Medical Sciences Romolo Laurita 1 , Anna Miserocchi 2 , Martina Ghetti 3 , Matteo Gherardi 1,2 , Augusto Stancampiano 4 , Valeria Purpura 3 , Davide Melandri 3 , Paola Minghetti 3 , Elena Bondioli 3 , Vittorio Colombo 1,2 Abstract— Sterilization of equipment and tissues is a common clinical practice: there are different chemical, mechanical and electromagnetic aseptic techniques for inactivating microorganisms. In particular, Skin Tissue Banks are investigating new methods to efficiently decolonize skin tissues, while preserving their structural features. In recent years, Cold Atmospheric Plasma (CAP) has demonstrated bactericidal, virucidal and fungicidal properties, due to the generation of reactive species and charged particles. For this reason, the aim of the present work is to demonstrate that the implementation of a Dielectric Barrier Discharge (DBD) treatment in air can effectively decontaminate skin tissue from Staphylococcus aureus, retaining cell viability and skin integrity. Fresh skin samples, taken from multi-tissue donors, were contaminated with Staphylococcus aureus and treated with a DBD source, to verify the level of bacterial decontamination induced by plasma. Cell viability and structural properties of skin tissue were investigated using MTT assay and hematoxylin-eosin staining, respectively. Our results show that CAP can sterilize skin tissue with a bacterial load up to 10 3 CFU/cm 2 ; moreover, it does not affect cell viability, and no loss of skin structural properties was observed. Thus, CAP treatment could be considered an innovative method for decolonization of human skin, without inducing any microscopic tissue damage, while keeping good cell viability. Index Terms— bacterial decontamination, cold atmospheric-plasma treatment, dielectric barrier discharge, plasma medicine, skin integrity. I.INTRODUCTION Sterilization of equipment and tissues is a common and necessary clinical practice and it is very important in a variety of applications, ranging from in vitro assays to in vivo treatment of wounds and skin grafts. There is a wide range of different chemical, physical and electromagnetic techniques to provide decontamination and sterilization [1]. Disinfection of skin allografts is mandatory, but decontamination standards have not been identified yet [2]. The use of allografts to replace damaged skin represents an important therapeutic resource, but the risk of transmitting infectious diseases has to be minimized, since 1 Department of Industrial Engineering, Alma Mater Studiorum - Università di Bologna, Italy 2 Industrial Research Centre for Advanced Mechanics and Materials, Alma Mater Studiorum - Università di Bologna, Italy naturally occurring contamination can be present on skin or can be introduced during processing [3]. Staphylococcus aureus, a pyogenic microorganism that normally resides on the human body as a commensal bacterium, has been identified as one of the most important pathogen responsible for skin and soft tissue infections [4] and can lead to nosocomial infections such as pneumonia and sepsis [5]. Moreover, S. aureus is one of the pathogens most frequently found after a donor explant, when grafts are delivered at the Skin Bank and are subjected to sterility analysis [3]. It causes numerous infections and possesses the ability to spread from person to person, by evading the immune response and acquiring resistance to antibiotics [6]. Antimicrobial-resistant infections represent a growing threat that the world faces today, since it causes at least 50,000 death per year in Europe and the United States. A constant rise in resistance by 2050 is estimated to lead to 10 million deaths per year worldwide and to a reduction of about of 2% of the gross domestic product. [7] The potential future annual costs related to the antimicrobial resistance are estimated to be about the 3.1% of global gross domestic product [8]. According to usual clinical best practice of the Regional Skin Bank “M. Bufalini”, the bacterial load of skin samples after explant is evaluated after 24 h of incubations on agar plates. If the bacterial load is higher than 10 2 CFU/cm 2 , donor tissues are consequently discarded according to the guidelines of the Italian National Transplant Centre [9], since conventional treatment (such as antibiotics) does not guarantee a total sterility and could cause onset of resistance in bacteria. In fact, sterility is the most important condition for an allograft, in order to avoid severe complications in deep or widespread burn patients, already highly weakened for the loss of integrity of the skin barrier. Secondly, during tissue processing and storage, it is very important to preserve cell viability and structural properties of the skin allograft, since it provides a temporary barrier against infection and fluid loss, it decreases pain and it promotes wound healing through the release of bioactive factors [10]. For all these reasons Skin Banks [11] are exploring novel and innovative methods to sterilize, or, at least, strongly reduce the initial bacterial load. One of these methods consists in the application of cold atmospheric plasmas (CAPs) for the sterilization of tissues, that could avoid the waste of contaminated donor tissue [12]–[16]. In recent years, CAPs are receiving a growing interest, mostly for their ability to inactivate viruses, bacteria and fungi, due to 3 Burn Centre and Emilia Romagna Regional Skin Bank, “M. Bufalini” Hospital, Cesena, Italy 4 Department for Life Quality Studies Alma Mater Studiorum - Università di Bologna, Italy. Cold atmospheric plasma treatment of infected skin tissue: evaluation of sterility, viability and integrity