The influence of different preservation and sterilisation steps on the histological properties of amnion allografts--light and scanning electron microscopic studies

Abstract

Despite thorough donor screening and preparation under aseptic conditions, conventional methods of preservation do not exclude the probability of a contamination with pathogenic germs. The purpose of this study was to investigate the changes of histological parameters of amnion transplants (ATs) through different methods of sterilisation and preservation. Therefore 10 different procedures for sterilisation and preservation of ATs were described. Specimens of each group were studied using different histological procedures such as light microscopy and scanning electron microscopy. General staining (Haematoxylin-eosin stain, periodic-acid-Schiff, Domack) and immunohistochemical methods have been applied in order to gain additional information concerning the structure of the amniotic epithelium and the basement membrane but also the distribution of collagens and intermediate filaments. Furthermore, the measurement of the ATs thickness was included in order to study the influence of the manufacturing procedures to this property. As a result we found that the histological appearance of the ATs is closely related to the applied sterilisation and preservation procedures. Although the basement membrane remained intact, especially the amniotic epithelium was partially destroyed by irradiation sterilisation. Further, the dissolution of the connective tissue layers into single fibre bundles was clearly visible. Procedures with and without peracetic acid sterilisation (PAA) preserved the tissue structure. Our results showed a significant variation in the tissue's thickness after different preservation procedures. Air- and freeze-dried ATs were found to be the thinnest tissues varying from 20 to 30 microm, the thickest ATs preserved in glycerol varied from 45 to 50 microm. Because ATs showed a preserved tissue structure after PAA sterilisation it can be recommended as an alternative for methods previously described in literature. Depending on the specific use of the AT one may choose from thinner or thicker allografts.