Establishment of an efficient method to decontaminate, wash and freeze cardiovascular tissues

Purpose: Establish a method to decontaminate, wash and freeze cardiovascular tissues.

Methods: Samples sterile porcine cardiovascular tissues, decontaminationantibioticsolutionBASE128_125ml/g 14h 37°C, washing: BASE 6.5 ml/g. 2×5 min. 1x6h. 4°C, RESEP treatment of hormogenates (antibiotic removal device): 2×20 min. RT.

Evaluation of:
1. Decontamination efficiency: samples were immersed in contaminated solutions (103-108 cfu/ml: 10 bacteria and 2 fungi strains), decontaminated, washed and homogenised. Homogenates (RESEP treated/untreated) were tested for contamination on agar plates and BactAlert.
2. Capability of washing procedure to remove antibiobiotic: decontamination and washing procedure was applied. 12 microbial strains were spiked on RESEP treated/untreated homogenates, tested on agar plates and BactAlert.
3. Freezing method: several profiles were tested on aortic, pulmonary and pericardium specimens.

Results:
1. Decontamination resulted in 6 Log 10 reduction for 5 strains (P.aeruginosa, B.atrophaeus, S.marcescens, P.acnes, E.coli), 5 Log 10 for 4 strains (S.epidermidis, K.pneumoniae, C.sporogenes, B.fragilis), 4 Log 10 for S.pyogenes, 1.0 Log 10 for C.albicans and A.brasiliensis.
2. Microbial growth was demonstrated on:
– RESEP-treated tissues: 12/12 strains (agar plates and BactAlert)
– RESEP-untreated tissues: 11/12 strains on agar plates (at less extent for 4 strains), 10/12 strains on BactAlert.
3. A freezing profile was defined leading to optimal freezing curve (-1°C/min to -40°C, -3°C/min to -120°C).

Conclusions: Decontamination was effective (4-6 Log 10) for 10/12 strains. The extensive washing procedure, able to remove antibiotic from the solutions and tissues. In combination with the RESEP ensures a reliable sterility assay. An optimal freezing profile was set-up.