Biologicals (2001) 29, 59–66 doi:10.1006/biol.2001.0286, available online at http://www.idealibrary.com on Validation of the Sterilization Procedure of Allogeneic Avital Bone Transplants Using Peracetic Acid–Ethanol Axel Pruss 1 , Ba ¨ rbel Baumann 2 , Michael Seibold 3 , Moujahed Kao 1,3 , Kathrin Tintelnot 3 , Ru ¨ diger von Versen 4 , Hartmut Radtke 1 , Thomas Do ¨ rner 5 , Georg Pauli 3 * and Ulf Berthold Go ¨ bel 2 1 Institute for Transfusion Medicine, Tissue Bank, University Hospital Charite ´, Medical Faculty of Humboldt-University, Berlin, Germany; 2 Institute for Microbiology and Hygiene, University Hospital Charite ´ , Medical Faculty of Humboldt-University, Berlin, Germany; 3 Robert Koch-Institut, Berlin, Germany; 4 German Institute for Cell and Tissue Replacement, Berlin, Germany; 5 Medical Clinic Centre Rheumatology and Clinical Immunology, University Hospital Charite ´, Medical Faculty of Humboldt-University, Berlin, Germany Abstract. Different procedures are available to inactivate bacteria and fungi, including their spores, as well as viruses in human bone transplants. The most efficient methods are considered to be gamma irradiation and thermal inactivation as well as chemical sterilization methods like the peracetic acid–ethanol treatment (PES). Following national and international standards or draft standards, the antimicrobial effectiveness of this procedure was evaluated. Due to the standardizable size as well as the clinical relevance, defatted human spongiosa cuboids (15×15×15 mm) served as model system. After treatment with PES for 2 and 4 hours, respectively, the titre of living micro-organisms was determined in the supernatant and the cuboid. A reduction in the titre of viable micro-organisms below the detection level (reduction factor >5 log 10 ) was already achieved after an incubation time of 2 hours (Staphylococcus aureus, Enterococcus faecium, Pseudomonas aeruginosa, Bacillus subtilis, Clostridium sporogenes, Mycobacterium terrae, Candida albicans as well as spores of Bacillus subtilis). No viable micro-organisms could be detected in any of the PES-treated test cuboids. Spores of Aspergillus niger were also completely inactivated. The PES pro- cedure proved to be a reliable method for the sterilization of human bone transplants derived from spongiosa. © 2001 The International Association for Biologicals Key words: sterilization, bone transplants, spongiosa, peracetic acid. Introduction In Germany about 25 000 allogeneic bone graft transplantations are performed annually. 1 Human bone tissue serves as source material for the produc- tion of these transplants (among others heads of femur and tibia, body of vertebra, iliac crest). Accordingly, special requirements have to be made on the biological safety of the tissues. Apart from a validated clinical e#ectiveness of the transplants, the quality assurance measures must focus on the validation of the sterilization process. In accord- ance with the guidelines for managing a bone tissue bank issued by the Bundesa ¨ rztekammer (Federal Medical Society), 2 sterilization is particularly indi- cated if a second testing of the bone tissue donor after an appropriate time cannot be performed, because the tissue was obtained from a cadaveric source. In addition, these validations are required standards in Germany, where all transplants manufactured in national tissue banks are regarded as medicinal products. So far, no normative specifications exist regard- ing validation of the disinfection or sterilization of bone tissue intended to be used for transplan- tation. Table 1 shows a survey of standards or draft standards, on which the design of validation experiments ought to be based. The aim of the projected validation design was to follow the relevant above-mentioned standards as *To whom correspondence should be addressed. E-mail: paulig@rki.de 1045–1056/01/020059+08 $35.00/0  2001 The International Association for Biologicals