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CD34+ cell selection from frozen cord blood products using the Isolex 300i and CliniMACS CD34 selection devices
I K McNiece1, G B Stoney, B P Kern
1Department of Developmental Hematology, Amgen Inc., Thousand Oaks, CA 91320, USA.
Insights
Clinical scale devices effectively select CD34+ cells from frozen cord blood (CB) for ex vivo expansion. The CliniMACS system demonstrated superior recovery of these essential progenitor cells compared to the Isolex 300i.
Area of Science:
- Hematology
- Cell Biology
- Biotechnology
Background:
- Ex vivo expansion of cord blood (CB) cells necessitates CD34+ cell selection for optimal progenitor cell yields.
- Preclinical development requires efficient methods for isolating CD34+ cells from cryopreserved CB products.
- Stem cell depletion during expansion necessitates using only a fraction of CB for expansion, preserving the remainder unmanipulated.
Purpose of the Study:
- To evaluate two clinical-scale cell selection devices, Isolex 300i and CliniMACS, for CD34+ cell selection from frozen CB.
- To assess the purity and recovery rates of CD34+ cells obtained using both devices.
- To determine the feasibility of using these devices for preclinical development of CB expansion strategies.
Main Methods:
- Frozen CB products were thawed, and 40% fractions were processed using either the Isolex 300i or CliniMACS device.
- Viability and total white blood cell counts were assessed post-thaw.
- Purity and recovery of CD34+ cells were determined for each device.
- Expanded CD34+ cells were analyzed for colony-forming unit-granulocyte-macrophage (CFU-GM) expansion.
Main Results:
- Both devices processed thawed CB fractions with >95% cell viability and a median WBC count of 1.8 x 10^8.
- The Isolex 300i yielded a median CD34+ cell purity of 51% and recovery of 34%.
- The CliniMACS system achieved a median CD34+ cell purity of 54% and a significantly higher recovery of 80%.
- Expansion of CD34+ cells from both systems resulted in >20-fold CFU-GM expansion, with a median of 44-fold.
Conclusions:
- Both Isolex 300i and CliniMACS are feasible for clinical-scale CD34+ cell selection from frozen CB products.
- The CliniMACS system offers superior recovery of CD34+ cells compared to the Isolex 300i.
- These findings support the use of clinical-scale selection devices for preparing CB for ex vivo expansion and potential therapeutic applications.
Abstract:
Ex vivo expansion of cord blood (CB) cells requires CD34+ cell selection before expansion to obtain optimal numbers of progenitor cells. As a preliminary step to preclinical development of CB expansion, we have evaluated two clinical scale selection devices, the Isolex 300i (Baxter Healthcare, Immunotherapy Division) and the CliniMACS (Miltenyi Biotech Inc.), for CD34+ cell selection from frozen CB products. As expansion of CB results in differentiation of cells, there may be a depletion of stem cells. Therefore, only a fraction of the CB should be expanded while a portion of the CB is maintained unmanipulated for infusion. After thawing of 40% fractions of each CB product, we observed >95% viable cells, with a median total WBC count of 1.8 x 10(8) cells. Use of the Isolex 300i resulted in a median purity of 51% CD34+ cells (n=8) and a median recovery of 34% CD34+ cells. Use of the CliniMACS resulted in a median purity of 54% CD34+ cells (n=10) and a median recovery of 80% CD34+ cells. The absolute number of CD34+ cells recovered after selection varied with samples from 6.7 x 10(4) to 3.2 x 10(6) CD34+ cells. Expansion of CD34+ cells from both systems resulted in >20-fold expansion of CFU-GM, with a median of 44-fold expansion. These data demonstrate the feasibility of selecting small fractions of frozen CB products using clinical scale CD34+ cell selection devices.