Waste Not, Want Not – Culturing CD34+ Hematopoietic Stem Cells from Used Leukoreduction Filters
A leukoreduction filter is flushed with buffer to harvest blood cells, including CD34+ hematopoietic stem cells. Image source: Neron, S. et al [2].

Most routine blood donations in the US are sifted through leukoreduction systems to offload their white blood cells. A new protocol featuring BioCision’s CoolCell® freezing containers details how to harvest CD34+ hematopoietic stem cells from the material trapped in those filtering devices.

According to the American Red Cross, 13.6 million blood donations are collected in the US every year. The sanguine fluid is sorely needed – 16 million transfusions are routinely performed in American hospitals, and supply is lagging behind demand. Before the donated blood reaches a patient, it is routinely tested for infectious diseases such as HIV and hepatitis B and C, and often passed through leukoreduction systems to deplete it of its white blood cells. Those white blood cells have a higher concentration of residual pathogens and may also interact with the immune system of the blood recipient – so removal of them significantly reduces post-transfusion infection rates.

Once a leukoreduction system has done its work, it is usually discarded. That is a crying shame, according to authors of a recent publication that outlines a safe, standardized method to harvest valuable stem cells from those discarded leukoreduction units. [1] They point out that each of those instruments traps millions of viable CD34+ hematopoietic stem cells during routine application, in much higher concentration than found directly in the human body. Talk about precious waste.

The authors’ protocol is surprisingly simple and starts with an easy manual washing procedure. The leukoreduction device is flushed with a buffer that contains a balanced salt solution (HBSS) and 10% anticoagulant citrate-dextrose (ACD). Subsequently, the obtained cells are concentrated via centrifugation in a Ficoll density gradient, and then cryopreserved in serum-free medium. To ensure maximal cell survival during the freezing process and optimal post-thaw viability and proliferative activity, the researchers recommend the use of BioCision’s CoolCell® freezing containers for the cryopreservation step.

The scientist continue to describe an immunomagnetic purification system that enriches the desired CD34+ hematopoietic stem cells. Cells are then expanded for 7 – 10 days in a custom-designed culture medium that is free of any animal protein and contains a cytokine cocktail that suppresses early progression of the CD34+ hematopoietic stem cells into the erythroid lineage. The authors finally explain flow cytometry approaches to characterize the diversity of the obtained cellular product, using antibodies and fluorochromes.

Standardization of these methods is necessary to ensure that the results of such enrichment procedures are of acceptable quality to be employed in downstream cellular therapies. Calls for such standardization have been gaining traction and support for some time (see one of our previous blogs). Publication of reliable, easily implementable protocols, including guidelines on cryopreservation techniques, fosters the development of a framework that increases safety, and, with that, public acceptance, of cellular therapies as a powerful tool in our arsenal against serious diseases.

And that, we think, is bloody brilliant.

References

[1] Cloutier, M. et al. In Vitro Culture of Human Hematopoietic Stem Cells in Serum Free Medium and Their Monitoring by Flow Cytometry. Methods Mol Biol 2016 Apr 1. [Epub ahead of print] PMID:27032946

[2] Néron, S. et al. Characterization of mononuclear cells remaining in the leukoreduction system chambers of apheresis instruments after routine platelet collection: a new source of viable human blood cells. Transfusion 47(6):1042-9. 2007.