It is well-known that cornea transplantation is still associated with a certain – though quite low –risk of post keratoplasty infection that can pose serious threats to the transplanted patient. Since one of the possible causes includes the grafting of contaminated donor tissue, the eye banks have the responsibility to ascertain that only corneas that are free of contamination are distributed for the transplantation.

With this aim, the Alchilife R&D laboratories have an ongoing collaboration with the Eye Bank of Monza to select and validate a system to confirm the sterility of corneas by testing all the media that are used to preserve and transport the donor tissues.

As we know, most of the European eye banks store the donor corneas at 31°C in organ culture media allowing to preserve donor tissues for up to 4 weeks. This longer preservation time gives the eye banks operators the ease of performing all accurate tests on donated tissues.

The corneal storage media available in the market contain a mixture of antimicrobial agents. On one side these agents help to minimize the contamination in the preserved corneas, but on the other, they may prevent the growth of some microorganisms whose presence might not be revealed during the microbiological analysis and, as a consequence, transferred to the transplanted patient.

This is the reason why the European Pharmacopeia (EP) “Method suitability test” recommends to remove all factors that may interfere with the microbial growth before the microbiological testing.

The same Eye Bank of Monza usually performed the microbiological tests using the automatic BACTEC™ (Becton Dickinson) system, through which the preservation media samples are tested using specific BACTEC™ bottles containing resins that should maintain the antimicrobial residues. Despite the use of an automatic system, it has been shown that sometimes the results obtained can be misleading.

To avoid false-negative results, the new studied method includes an additional step in which all media to be tested are treated with a patented syringe-like CE-marked medical device, RESEP (Alchimia Srl), which also contains a special resin mixture for total elimination of the antimicrobials from the corneal storage media.

This method was validated using both samples of organ culture medium (Tissue-C, Alchimia Srl) and deswelling/transport medium (Carry-C, Alchimia Srl) by inoculum with various EP reference microbial strains before the transfer into the BACTEC™ bottles for the automatic analysis. Of course, negative controls (i.e. samples without microbial strains) and positive controls (i.e. samples with microbial strains) were also used.

The obtained results showed that the use of RESEP increased the sensitivity of the sterility test using BACTEC™ automated system up to 100% and, consequently, allowed validation of the method for sterility testing of corneal storage and deswelling/transport media according to the EP requirements.

References:

  • Vignola R. et al. Validation of the BD BACTEC™ method for sterility testing of corneal preservation media according to the European Pharmacopoeia (chapter 2.6.1.). Presentations at the European Association of Tissue Banks EATB 2017 and 2018
  • Vignola et al. A two-centre validation study of sterility test of corneal storage media with elimination of interfering antimicrobials in compliance with the European Pharmacopoeia. Cell Tissue Bank 2019 https://doi.org/10.1007/s10561-019-09766-7
  • Council of Europe (2013a) Sterility test. In: European Directorate For The Quality of Medicines and Healthcare (ed) European Pharmacopoeia, Eight edn, Vol I, Strasbourg Cedex, France, pp 175–178
  • Schroeter J. et al. Validation of the microbiological testing of tissue preparations using the BACTECTM blood culture system. Transfus Med Hemother 2012;39(6):387–390
  • Mistò R. et al. Method for sterility testing of corneal storage and transport media after removal of interfering antimicrobials: prospective validation study in compliance with the European Pharmacopoeia. BMJ Open Ophthalmol 2017;2(1): e000093.