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An objective flow cytometry method to rapidly determine cord blood potency in cryopreserved units
Author(s) -
Simard Carl,
Bonnaure Guillaume,
Fournier Diane,
Néron Sonia
Publication year - 2019
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1111/trf.15220
Subject(s) - cryopreservation , cord blood , flow cytometry , andrology , potency , cd34 , chemistry , immunology , biology , medicine , stem cell , in vitro , biochemistry , microbiology and biotechnology , embryo
BACKGROUND Cord blood banks have to determine the regenerative potential of cord blood units (CBUs) on a representative sample of the cryopreserved product before release to the transplant center. Potency can be measured by using a colony‐forming unit (CFU) method, which delays the release of CBU by 7 to 14 days. To accelerate CBU qualification, we have developed a rapid method to assess the response of CD34 cells to interleukin (IL)‐3. Flow cytometry was used to measure IL‐3–induced STAT5 phosphorylation within CD34‐cells. This IL‐3 test was compared to the CFU method, as well as the aldehyde dehydrogenase (ALDH) enzyme‐based assay. STUDY DESIGN AND METHODS Ten cryopreserved CBUs were analyzed for their contents in CD34 and CD45 viable cells, total CFUs, ADLH bright cells, and IL‐3–responsive CD34+ cells. Extreme and mild warming event scenarios were simulated on CBUs and used as poor‐quality samples. Segments, tubes, and bags from five CBUs were compared for their potency using IL‐3 and CFU methods. RESULTS The IL‐3 test was accurate in identifying the samples handled following standard operating procedures and those subjected to extreme warming events. Based on these results, a threshold of 55% of IL‐3–responsive CD34 cells was established to identify good‐quality samples. The IL‐3 test was also the most sensitive to detect samples subjected to milder warming events. CONCLUSIONS Our new method for determining CBU functionality is rapid, unbiased, and robust. The IL‐3 test described herein fulfills the requirements for validation, and we intend to implement this method in our cord blood bank facility.