Processing Storage And Preparation For

In 1995, the National Heart, Lung, and Blood Institute funded three UCB banks and six transplant centers to assist in establishing standard operation procedures for the collection, processing, and investigation of UCB use.14 Several modifications of the standard procedure of UCB processing and storage have been developed to reduce unit size (and thus storage costs) and enhance red blood cell (RBC) depletion while maintaining sterility and HSC composition and activity.20-26 Three to five milliliters of UCB is removed for HLA-typing, disease testing, microbiological cultures, progenitor cell assays, and assessment of total nucleated and CD34+ cell counts. RBC depletion and volume reduction begin with the addition of 6% hydroxyethyl starch to enhance RBC sedimentation. The mixture is centrifuged at 50g for 5 minutes. The leukocyte-containing supernatant is then centrifuged at 400g for 10 min yielding a sediment of white cells. The leukocyte pellet is resuspended in plasma and diluted to a final volume of 20 ml.

UCB is cryopreserved with chilled dimethyl sulfox-ide to a final concentration of 10% and a total volume of 25 ml. The sample is placed in aluminum containers and frozen to -50°C in a -80°C freezer. The UCB unit is then transferred to a liquid nitrogen storage device where it may be housed for several years.27

Preparation of a frozen UCB unit for infusion is similar to the procedure used for PB stem cell units. The unit is placed in a sterile, sealable plastic bag, submerged, and gently agitated in a 37°C water bath. When thawed, equal volumes of 10% dextran and 5% albumin are added to a volume double of that was originally collected. The suspension is then cen-trifuged at 250g for 10 min. The pellet is resuspended with equal volumes (50 ml) of 10% dextran and 5% albumin. Samples of the final unit are sent for bacterial and fungal cultures.

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