Evaluation of Alternative Samples for Sterility Testing of HPC-CB Products

Background: The infused cell dose is an important determinant of engraftment after CB transplantation. Thus, cell loses during manufacturing and samples for testing must be minimized. The AutoXpress® (AXP) system is used for processing of HPC-CB FDA licensed products in our Bank. With this, collected CB is separated into three components (fractions): red blood cells (RBC), plasma (P) and mononuclear cells (MNC), which represent the final HPC-CB product.  Methods: to optimize the number of HPC-CB cells to be cryopreserved, we explored the suitability of RBC or P samples as surrogates of the final product for testing of bacteria/fungi contamination.   We evaluated the distribution of 7 microorganisms, spiked into whole CB collections at known concentrations (triplicate studies for each microorganism), into each of the three post-processing fractions.  Further, a Rapid Microbiological Method (VersaTREK, ThermoFisher Scientific, OH) was used for the detection of bacteria/fungi in samples of RBC, MNC and P and compared to the gold standard bacteriology method (USP<71>). Results: Spiked microorganisms were distributed differently: Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus epidermidis, Propionibacterium acnes (Pac) and Bacillus subtilis were concentrated preferentially in the MNC, while Candida albicans and Aspergillus Niger were more so in the RBC fraction. Nonetheless, the VersaTREK detected all microorganisms (sensitivity ≤100 CFU/ bottle) from all three fractions with a comparable time to detection (TTD) for RBC and MNC samples (all ≤ 14 days). A 10-fold higher P volume (5 mL) was used to detect growth at a TTD comparable to MNC or RBC samples. Further, delayed detection, due to partial inhibition of the growth of Pac, was seen with the higher P volume. 

Summary: The VersaTREK/Redox detects microorganisms with sensitivity comparable to the standard method in all three fractions after AXP-processing. Use of MNC or RBC did not impact significantly the ability or time to detection. The use of samples from the RBC or P fractions, or possibly, a combination of both for sterility screening of HPC-CB products will preserve approx. 5% of the final product volume for transplant use.