StemSpan™ SFEM

Serum-free medium for culture and expansion of hematopoietic cells

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Serum-free medium for culture and expansion of hematopoietic cells
From: 116 USD

Overview

StemSpan™ Serum-Free Expansion Medium (SFEM) has been developed and tested for the in vitro culture and expansion of human hematopoietic cells, when the appropriate growth factors and supplements are added. This allows users the flexibility to prepare medium that meets their requirements. When combined with the appropriate cytokines, SFEM has been used for the culture and expansion of hematopoietic cells isolated from other species, including mouse, non-human primate, and dog. SFEM has also been used for culture of various other hematopoietic and non-hematopoietic cell types. Using appropriate StemSpan™ Expansion Supplements, SFEM 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 progenitors to generate populations of erythroid, myeloid, or megakaryocyte progenitor cells.
Contains:
• Iscove’s MDM
• Bovine serum albumin
• Recombinant human insulin
• Human transferrin (iron-saturated)
• 2-Mercaptoethanol
• Supplements
Subtype:
Specialized Media
Cell Type:
Hematopoietic Stem and Progenitor Cells
Species:
Human; Mouse; Rat; Non-Human Primate
Application:
Cell Culture; Expansion
Brand:
StemSpan
Area of Interest:
Stem Cell Biology; Transplantation Research
Formulation:
Defined; Serum-Free

Scientific Resources

Product Documentation

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Educational Materials

(9)

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

Expansion of CD34 + Human Cord Blood Cells Cultured in StemSpan™ Media Containing CC100 Cytokine Cocktail

Figure 1. Expansion of CD34 + Human Cord Blood Cells Cultured in StemSpan™ Media Containing CC100 Cytokine Cocktail

Purified CD34 + human cord blood (CB) cells were suspended at a concentration of 10,000 per mL in StemSpan™ SFEM (dark gray bars), SFEM II (gold bars) and ACF (orange bars) media containing CC100 Cytokine Cocktail (Catalog #02690). Cultures were maintained for 7 days, after which the cells were counted and examined for CD34 and CD45 expression by flow cytometry. Shown are the fold expansion of total nucleated cells (TNC) (A) and CD34 + cells (B) per input CD34 + cell, and the percent CD34 + cells (C). Results represent the average results of 32 different CB samples. Vertical lines indicate 95% confidence limits, the range within which 95% of results fall. The numbers of cells produced in StemSpan™ SFEM II were significantly higher than in StemSpan™ SFEM and StemSpan™-ACF (*p<0.001, paired t-test, n=32).

StemSpan™ SFEM II Serum-Free Expansion Medium Containing CC100 Cytokine Cocktail Supports Greater Expansion of Human CD34 + Cells Than Other Media Tested

Figure 2. StemSpan™ SFEM II Serum-Free Expansion Medium Containing CC100 Cytokine Cocktail Supports Greater Expansion of Human CD34 + Cells Than Other Media Tested

Expansion of CD34 + cells, normalized relative to the values obtained in StemSpan™ SFEM medium (dark gray bars) after culturing purified CD34 + CB (A, n=6) or bone marrow (BM) (B, n=3) cells for 7 days in StemSpan™ SFEM, SFEM II (gold bars) and ACF (orange bars), and six media from other commercial suppliers (light gray bars, Competitor 1-6, which included, in random order, StemPro34 (Life Technologies), X-Vivo-15 and HPGM (both from Lonza), SCGM (Cellgenix), StemLine II (Sigma) and HP01 (Macopharma)). All media were supplemented with StemSpan™ CC100 Cytokine Cocktail (Catalog #02690). Vertical lines indicate 95% confidence limits, the range within which 95% of results fall. The numbers of CB and BM cells produced in StemSpan™ SFEM II were significantly higher than in all other media, except the numbers of CB cells produced in StemSpan™-ACF (*p<0.05, paired t-test).

Expansion of CD34 + Human Cord Blood Cells Cultured in StemSpan™ Media Containing CD34 + Expansion Supplement

Figure 3. Expansion of CD34 + Human Cord Blood Cells Cultured in StemSpan™ Media Containing CD34 + Expansion Supplement

Purified CD34 + human cord blood (CB) cells were suspended at a concentration of 10,000 per mL in StemSpan™ SFEM (dark gray bars), SFEM II (gold bars) and ACF (orange bars) media containing CD34 + Expansion Supplement (Catalog #02691). Cultures were maintained for 7 days, after which the cells were counted and examined for CD34 and CD45 expression by flow cytometry. The number of colony-forming units (CFU) in the expanded population was determined by replating cells in MethoCult™ H4435 and counting the number of colonies produced 14 days later. Shown are the fold expansion of total nucleated cells (TNC) (A), CD34 + cells (B) and CFU numbers (C) per input CD34 + cell, and the percent CD34 + cells (D) in these cultures (n=6). Vertical lines indicate 95% confidence limits, the range within which 95% of results fall. The numbers of cells produced in StemSpan™ SFEM II was significantly higher than in SFEM and ACF (*p<0.001, #p<0.05, paired t-test, n=6).

StemSpan™ SFEM II Serum-Free Expansion Medium Containing CD34 + Expansion Supplement Supports Greater Expansion of Human CD34 + Cells Than Other Media Tested

Figure 4. StemSpan™ SFEM II Serum-Free Expansion Medium Containing CD34 + Expansion Supplement Supports Greater Expansion of Human CD34 + Cells Than Other Media Tested

Expansion of CD34 + cells (A) and CFUs (B), normalized relative to the values obtained in SFEM medium (dark gray bars) after culturing purified CD34 + CB cells for 7 days in StemSpan™ SFEM, SFEM II (gold bars) and ACF (orange bars), and six media from other suppliers (light gray bars, Competitor 1-6, which included, in random order, X-Vivo-15 (Lonza), HP01 (Macopharma), StemPro34 (Life Technologies), SCGM (Cellgenix), StemLine II (Sigma), and HPGM (Lonza). All media were supplemented with the StemSpan™ CD34 + Expansion Supplement (Catalog #02691). Vertical lines indicate 95% confidence limits, the range within which 95% of results fall. The numbers of cells produced in StemSpan™ SFEM II were significantly higher than in all other media (*p<0.01, paired t-test, n=6).

Table 1. Production of Erythroid Cells From CD34 + Human Cord Blood Cells Cultured in StemSpan™ SFEM Serum-Free Expansion Medium Containing Erythroid Expansion Supplement

Production of Erythroid Cells From CD34 + Human Cord Blood Cells Cultured in StemSpan™ SFEM Serum-Free Expansion Medium Containing Erythroid Expansion Supplement

Numbers and percent of erythroid cells produced after 14 days of culture of enriched CD34 + cells from 14 different cord blood (CB) samples. Erythroid cells were characterized by flow cytometry on the basis of transferrin receptor (CD71) and glycophorin A (CD235) expression.*95% confidence limits, the range within which 95% of the results fall.

StemSpan™ SFEM II Serum-Free Expansion Medium Containing Erythroid Expansion Supplement Supports Greater Expansion of Erythroid Cells Than Other Media Tested

Figure 5. StemSpan™ SFEM II Serum-Free Expansion Medium Containing Erythroid Expansion Supplement Supports Greater Expansion of Erythroid Cells Than Other Media Tested

The numbers of erythroid cells, normalized relative to the values obtained in StemSpan™ SFEM medium (dark gray bar), obtained after culturing purified CD34 + CB cells for 14 days in StemSpan™ SFEM, SFEM II (gold bars) and ACF (orange bars), and six media from other commercial suppliers (light gray bars, Competitor 1-6, which included, in random order, X-Vivo-15 and HPGM (both from Lonza), StemLine II (Sigma), HP01 (Macopharma), StemPro34 (Life Technologies) and SCGM (Cellgenix). All media were supplemented with StemSpan™ Erythroid Expansion Supplement (Catalog #02692). Vertical lines indicate 95% confidence limits, the range within which 95% of results fall. The numbers of cells produced in StemSpan™ SFEM II were significantly higher than in all other media (*p<0.05, paired t-test, n=6).

Table 2. Production of Megakaryocytes From CD34+ Human Cord Blood Cells Cultured in StemSpan™ SFEM Serum-Free Expansion Medium Containing Megakaryocyte Expansion Supplement

Production of Megakaryocytes From CD34+ Human Cord Blood Cells Cultured in StemSpan™ SFEM Serum-Free Expansion Medium Containing Megakaryocyte Expansion Supplement

Numbers and percent of cells expressing the megakaryocyte marker CD41a produced after 14 days of culture of enriched CD34 + cells from 6 independent cord blood (CB) samples. *95% confidence limits, the range within which 95% of the results fall.

StemSpan™ SFEM II Serum-Free Expansion Medium Containing Megakaryocyte Expansion Supplement Supports Greater Expansion of Megakaryocytes Than Other Media Tested

Figure 6. StemSpan™ SFEM II Serum-Free Expansion Medium Containing Megakaryocyte Expansion Supplement Supports Greater Expansion of Megakaryocytes Than Other Media Tested

The numbers of megakaryocytes, normalized relative to the values obtained in StemSpan™ SFEM medium (dark gray bar), obtained after culturing purified CD34 + CB cells for 14 days in StemSpan™ SFEM, SFEM II (gold bars) and ACF (orange bars), and six media from other commercial suppliers (light gray bars, Competitor 1-6, which included, in random order, StemLine II (Sigma), HPGM (Lonza), HP01 (Macopharma), SCGM (Cellgenix), StemPro34 (Life Technologies) and X-Vivo-15 (Lonza). All media were supplemented with StemSpan™ Megakaryocyte Expansion Supplement (Catalog #02696). Vertical lines indicate 95% confidence limits, the range within which 95% of results fall. The numbers of cells produced in the StemSpan™ media were significantly higher than in the other media (*p<0.01 paired t-test, n=6).

Publications

(214)
Scientific reports 2018 APR

Efficient differentiation of cardiomyocytes and generation of calcium-sensor reporter lines from nonhuman primate iPSCs.

Y. Lin et al.

Abstract

Nonhuman primate (NHP) models are more predictive than rodent models for developing induced pluripotent stem cell (iPSC)-based cell therapy, but robust and reproducible NHP iPSC-cardiomyocyte differentiation protocols are lacking for cardiomyopathies research. We developed a method to differentiate integration-free rhesus macaque iPSCs (RhiPSCs) into cardiomyocytes with {\textgreater}85{\%} purity in 10 days, using fully chemically defined conditions. To enable visualization of intracellular calcium flux in beating cardiomyocytes, we used CRISPR/Cas9 to stably knock-in genetically encoded calcium indicators at the rhesus AAVS1 safe harbor locus. Rhesus cardiomyocytes derived by our stepwise differentiation method express signature cardiac markers and show normal electrochemical coupling. They are responsive to cardiorelevant drugs and can be successfully engrafted in a mouse myocardial infarction model. Our approach provides a powerful tool for generation of NHP iPSC-derived cardiomyocytes amenable to utilization in basic research and preclinical studies, including in vivo tissue regeneration models and drug screening.
Nature genetics 2018 APR

Natural regulatory mutations elevate the fetal globin gene via disruption of BCL11A or ZBTB7A binding.

G. E. Martyn et al.

Abstract

$\beta$-hemoglobinopathies such as sickle cell disease (SCD) and $\beta$-thalassemia result from mutations in the adult HBB ($\beta$-globin) gene. Reactivating the developmentally silenced fetal HBG1 and HBG2 ($\gamma$-globin) genes is a therapeutic goal for treating SCD and $\beta$-thalassemia 1 . Some forms of hereditary persistence of fetal hemoglobin (HPFH), a rare benign condition in which individuals express the $\gamma$-globin gene throughout adulthood, are caused by point mutations in the $\gamma$-globin gene promoter at regions residing {\~{}}115 and 200 bp upstream of the transcription start site. We found that the major fetal globin gene repressors BCL11A and ZBTB7A (also known as LRF) directly bound to the sites at -115 and -200 bp, respectively. Furthermore, introduction of naturally occurring HPFH-associated mutations into erythroid cells by CRISPR-Cas9 disrupted repressor binding and raised $\gamma$-globin gene expression. These findings clarify how these HPFH-associated mutations operate and demonstrate that BCL11A and ZBTB7A are major direct repressors of the fetal globin gene.
Nature communications 2017

An immortalized adult human erythroid line facilitates sustainable and scalable generation of functional red cells.

K. Trakarnsanga et al.

Abstract

With increasing worldwide demand for safe blood, there is much interest in generating red blood cells in vitro as an alternative clinical product. However, available methods for in vitro generation of red cells from adult and cord blood progenitors do not yet provide a sustainable supply, and current systems using pluripotent stem cells as progenitors do not generate viable red cells. We have taken an alternative approach, immortalizing early adult erythroblasts generating a stable line, which provides a continuous supply of red cells. The immortalized cells differentiate efficiently into mature, functional reticulocytes that can be isolated by filtration. Extensive characterization has not revealed any differences between these reticulocytes and in vitro-cultured adult reticulocytes functionally or at the molecular level, and importantly no aberrant protein expression. We demonstrate a feasible approach to the manufacture of red cells for clinical use from in vitro culture.
Nature 2017

Conversion of adult endothelium to immunocompetent haematopoietic stem cells.

Lis R et al.

Abstract

Developmental pathways that orchestrate the fleeting transition of endothelial cells into haematopoietic stem cells remain undefined. Here we demonstrate a tractable approach for fully reprogramming adult mouse endothelial cells to haematopoietic stem cells (rEC-HSCs) through transient expression of the transcription-factor-encoding genes Fosb, Gfi1, Runx1, and Spi1 (collectively denoted hereafter as FGRS) and vascular-niche-derived angiocrine factors. The induction phase (days 0-8) of conversion is initiated by expression of FGRS in mature endothelial cells, which results in endogenous Runx1 expression. During the specification phase (days 8-20), RUNX1(+) FGRS-transduced endothelial cells commit to a haematopoietic fate, yielding rEC-HSCs that no longer require FGRS expression. The vascular niche drives a robust self-renewal and expansion phase of rEC-HSCs (days 20-28). rEC-HSCs have a transcriptome and long-term self-renewal capacity similar to those of adult haematopoietic stem cells, and can be used for clonal engraftment and serial primary and secondary multi-lineage reconstitution, including antigen-dependent adaptive immune function. Inhibition of TGFβ and CXCR7 or activation of BMP and CXCR4 signalling enhanced generation of rEC-HSCs. Pluripotency-independent conversion of endothelial cells into autologous authentic engraftable haematopoietic stem cells could aid treatment of haematological disorders.
Science translational medicine 2016 OCT

Selection-free genome editing of the sickle mutation in human adult hematopoietic stem/progenitor cells.

M. A. DeWitt et al.

Abstract

Genetic diseases of blood cells are prime candidates for treatment through ex vivo gene editing of CD34+ hematopoietic stem/progenitor cells (HSPCs), and a variety of technologies have been proposed to treat these disorders. Sickle cell disease (SCD) is a recessive genetic disorder caused by a single-nucleotide polymorphism in the $\beta$-globin gene (HBB). Sickle hemoglobin damages erythrocytes, causing vasoocclusion, severe pain, progressive organ damage, and premature death. We optimize design and delivery parameters of a ribonucleoprotein (RNP) complex comprising Cas9 protein and unmodified single guide RNA, together with a single-stranded DNA oligonucleotide donor (ssODN), to enable efficient replacement of the SCD mutation in human HSPCs. Corrected HSPCs from SCD patients produced less sickle hemoglobin RNA and protein and correspondingly increased wild-type hemoglobin when differentiated into erythroblasts. When engrafted into immunocompromised mice, ex vivo treated human HSPCs maintain SCD gene edits throughout 16 weeks at a level likely to have clinical benefit. These results demonstrate that an accessible approach combining Cas9 RNP with an ssODN can mediate efficient HSPC genome editing, enables investigator-led exploration of gene editing reagents in primary hematopoietic stem cells, and suggests a path toward the development of new gene editing treatments for SCD and other hematopoietic diseases.
STEMCELL TECHNOLOGIES INC.’S QUALITY MANAGEMENT SYSTEM IS CERTIFIED TO ISO 13485. PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED.