Overview
Using appropriate cytokines, StemSpan™-ACF Without Phenol Red may be used to expand CD34+ cells isolated from human cord blood, mobilized peripheral blood, or bone marrow samples, or to expand and differentiate lineage-committed progenitor cells to generate populations of myeloid or megakaryocyte progenitor cells.
StemSpan™-ACF Without Phenol Red is manufactured and tested following relevant cGMPs under a certified quality management system. For additional quality information, visit www.STEMCELL.com/compliance.
⦁ Maintenance of primitive CD34brightCD90+CD45RA- population in culture
⦁ Suitable for use in genome editing protocols
⦁ Animal component- and phenol red-free formulation
⦁ Manufactured and tested under relevant cGMPs
⦁ Full traceability of raw materials
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(5)Product Applications
This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.
Data and Publications
Data

Figure 1. Day 7 Immunophenotyping of CD34+ Cells Cultured in StemSpan™-ACF Without Phenol Red
CD34+ cells were purified from cord blood (CB) using the EasySep™ Human Cord Blood CD34 Positive Selection Kit II (Catalog #17896) and cultured in StemSpan™-ACF Without Phenol Red (Catalog #100-0130) supplemented with StemSpan™ CD34+ Expansion Supplement (Catalog #02691) (A) without or (B) with the addition of UM729 (Catalog #72332). After 7 days, the cultured cells were stained with fluorescently labeled antibodies against CD34, CD90, and CD45RA, in addition to viability dye 7-AAD, and analyzed by flow cytometry. The horizontal dotted line in the CD34 vs FSC plots indicates the boundary between CD34- and CD34+ cells as based on a fluorochrome minus one (FMO) control for CD34 expression. Orange gates on these plots indicate the population of CD34bright cells used to generate data in Figures 2 and 3. Sequential gates were used to determine the percentages of viable CD34+ cells, CD34bright cells, and CD34brightCD90+CD45RA- cells.

Figure 2. StemSpan™ Media Support Greater Expansion of Human CD34+ and CD34bright Cells than Other Commercial Media
Purified CB-derived CD34+ cells were cultured for 7 days in select StemSpan™ media (StemSpan™ SFEM II, or StemSpan™-ACF, orange bars), and in five xeno-free media formulations from other suppliers (Xeno-Free Commercial Alternative, grey bars) including (in random order) CTS™ StemPro™ HSC (Thermo), SCGM (Cellgenix), X-VIVO™ 15 (Lonza), Stemline™ II (Sigma), and StemPro™-34 (Thermo). All media were supplemented with StemSpan™ CD34+ Expansion Supplement and UM171*. The (A) frequency and (B) cell expansion of viable CD34+ and CD34bright cells in culture were based on viable cell counts and flow cytometry results as shown in Figure 1. StemSpan™ SFEM II showed significantly higher expansion of CD34+ and CD34bright cells (P < 0.05 when comparing StemSpan™ SFEM II to five media from other suppliers, calculated using a one-way ANOVA followed by Dunnett’s post hoc test) and StemSpan™-ACF, the only animal component-free formulation, showed equivalent performance to all xeno-free competitors tested. Data shown are mean ± SEM (n = 8).
Note: Data for StemSpan™-ACF shown were generated with the original phenol red-containing version (Catalog #09855). However internal testing showed that the performance of the new phenol red-free cGMP-manufactured version of StemSpan-ACF(Catalog #100-0130) was very similar.
*Similar results are expected when using UM729 (Catalog #72332) prepared to a final concentration of 1μM. For more information including data comparing UM171 and UM729, see Fares et al., 2014.

Figure 3. StemSpan™ Media Support Greater Expansion of Human CD34+CD90+CD45RA- and CD34brightCD90+CD45RA- Cells than Other Commercial Media
Purified CB-derived CD34+ cells were cultured for 7 days in select StemSpan™ media (StemSpan™ SFEM II, or StemSpan™-ACF, orange bars), and in five xeno-free media formulations from other suppliers (Commercial Alternative, grey bars) including (in random order) CTS StemPro HSC (Thermo), SCGM (Cellgenix), X-VIVO 15 (Lonza), Stemline II (Sigma), and StemPro 34 (Thermo). All media were supplemented with StemSpan™ CD34+ Expansion Supplement and UM171*. The (A) frequency and (B) cell expansion of CD34+CD90+CD45RA- (solid) and CD34brightCD90+CD45RA- (dotted overlay) cells in culture were based on viable cell counts and flow cytometry results as shown in Figure 1. StemSpan™ media showed similar or significantly higher expansion of CD34brightCD90+CD45RA- cells (P < 0.05 compared to five media from other suppliers, calculated using one-way ANOVA followed by Dunnett’s post hoc test) and StemSpan™-ACF, the only animal component-free formulation tested, showed equivalent performance to all xeno-free competitors tested. Data shown are mean ± SEM (n = 8).
Note: Data for StemSpan™-ACF shown were generated with the original phenol red-containing version (Catalog #09855). However internal testing showed that the performance of the new phenol red-free cGMP-manufactured version of StemSpan-ACF(Catalog #100-0130) was very similar.
*Similar results are expected when using UM729 (Catalog #72332) prepared to a final concentration of 1μM. For more information including data comparing UM171 and UM729, see Fares et al. 2014.

Figure 4. StemSpan™ Media Support Better CD34+ and Primitive CD34+CD90+CD45RA- HSPC Expansion in a Genome Editing Application Compared with Alternative Commercial Media
Purified CB-derived CD34+ cells were cultured for 2 days in select StemSpan™ media (StemSpan™ SFEM II or StemSpan™-ACF, orange bars), or five xeno-free media formulations from other suppliers (gray bars). All media were supplemented with StemSpan™ CD34+ Expansion Supplement and UM171*. Cells were then electroporated using Arcitect™ CRISPR-Cas9 RNP complexes containing crRNA:tracrRNA targeting beta-2-microglobulin (B2M), and cultured for an additional 4 days in the same conditions. Knockout efficiency as measured by staining for MHC-I and analyzing by flow cytometry, was similar in all media tested, ~70-80%. (A) The percentage of CD34+ cells and (B) CD34+CD90+CD45RA-cells were quantified by flow cytometry 4 days post-electroporation. Data shown are mean + SD (n = 4 donors; **P < 0.01).
Note: Data for StemSpan™-ACF shown were generated with the original phenol red-containing version (Catalog #09855). However internal testing showed that the performance of the new phenol red-free cGMP-manufactured version of StemSpan-ACF Without Phenol Red (Catalog #100-0130) was very similar.
*Similar results are expected when using UM729 (Catalog #72332) prepared to a final concentration of 1 μM. For more information including data comparing UM171 and UM729, see Fares et al., 2014.