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StemSpan™

Hematopoietic Cell Expansion Media and Supplements

StemSpan™ serum-free hematopoietic cell expansion media promote the expansion and/or lineage-specific differentiation of human hematopoietic stem and progenitor cells (HSPCs) when supplemented with hematopoietic growth factors and/or other stimuli selected by the user. StemSpan™ Expansion Supplements are pre-mixed cocktails of recombinant human cytokines and other additives formulated to selectively promote the expansion of CD34+ stem and progenitor cells, or to stimulate their differentiation into erythroid, myeloid (granulocyte or monocyte) or megakaryocyte progenitor cells, when added to StemSpan™ media. A complete kit for the expansion and lineage-specific differentiation of CD34+ cells into T cells in stroma-free conditions is also available for research into lymphopoiesis.

Why Use StemSpan™ Media and Expansion Supplements?

  • Components are carefully selected to minimize lot-to-lot variability and support optimal and consistent culture results.
  • Media do not contain cytokines, allowing the flexibility to add StemSpan™ Expansion Supplements, cytokines and/or additives.
  • StemSpan™ SFEM II, combined with the appropriate Expansion Supplement, supports greater expansion of CD34+ cells and differentiation of erythroid cells, granulocytes, monocytes, megakaryocytes and T cells than other media tested.
  • StemSpan™-ACF is the first commercially available animal component-free medium for culturing HSPCs.

Expansion Media

StemSpan™ SFEM

StemSpan™ SFEM

Formulation:

Serum-free

Species:

  • Human
  • Mouse
  • Rat
  • Non-human primate

Recommended for:

Culture of HSPCs in serum-free conditions

Components:

  • BSA
  • Insulin
  • Tranferrin
  • Supplements
  • IMDM base

StemSpan™ SFEM II

StemSpan™ SFEM II

Formulation:

Serum-free

Species:

Human

Recommended for:

Culture of HSPCs in serum-free conditions

Components:

  • BSA
  • Insulin
  • Tranferrin
  • Supplements
  • IMDM base

StemSpan™ H3000

StemSpan™ H3000

Formulation:

Xeno-Free

Species:

Human

Recommended for:

Culture of HSPCs in the absence of non-human animal-derived components

Components:

  • Human-derived and recombinant human proteins
  • IMDM base

StemSpan™-ACF

Formulation:

Animal Component-Free

Species:

Human

Recommended for:

Culture of HSPCs in the absence of human- or animal-derived components

Components:

  • Recombinant and synthetic components
  • IMDM base
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Expansion Supplements for Human HSPCs

Expansion of CD34+ Cells

Application:

Culture and expansion of human CD34+ hematopoietic cells

Differentiation of Erythroid, Myeloid and Megakaryocyte Lineage Cells

Application:

Expansion and lineage-specific differentiation of human CD34+ cells to generate large numbers of erythroid progenitor cells, granulocytes, monocytes or megakaryocytes

Differentiation of CD34+ Cells into T Cells

Application:

Expansion and lineage-specific differentiation of cord-blood derived CD34+ cells to generate T cells in stroma-free conditions
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Data

Figure 1. 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 bar) after culturing purified CD34+ CB cells for 7 days in StemSpan™ SFEM, SFEM II (gold bar) and ACF (orange bar), 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™ CC100 Cytokine Cocktail. 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, except the numbers of CB cells produced in StemSpan™-ACF (*p<0.05, paired t-test, n=6).



Figure 2. 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 normalized relative to the values obtained in SFEM medium (dark gray bar) after culturing purified CD34+ CB cells for 7 days in StemSpan™ SFEM, SFEM II (gold bar) and ACF (orange bar), and six media from other 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 the StemSpan™ CD34+ Expansion Supplement. 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).



Figure 3. 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 bar) and ACF (orange bar), 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™ Erythroid Expansion Supplement. 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).



Figure 4. 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 bar) and ACF (orange bar), 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).



Table 1. Production of Myeloid Cells from Human CB CD34+ Cells Cultured in SFEM II Containing Myeloid Expansion Supplement or Myeloid Expansion Supplement II

Shown are numbers of total nucleated cells (TNCs) produced per input human CB-derived CD34+ cell and percentages of cells positive for myeloid markers CD13, CD14 and CD15 after 14 days of culture in SFEM II containing Myeloid Expansion Supplement (n = 14) or Myeloid Expansion Supplement II (n = 16). *95% confidence limits (CL); the range within which 95% of results typically fall.



Figure 5. Frequency and Yield of CD4 ISP and CD4+CD8+ DP Cells After 42 Days of Culture

CB-derived CD34+ cells (freshly isolated or frozen) were cultured with the StemSpan™ T Cell Generation Kit (Catalog #09940) for 42 days and (A) analysed by flow cytometry for the expression of CD4, CD8, CD3 and TCRαβ. The (B) frequency and (C) yield of CD4 ISP, double-positive (CD4+CD8+) and CD3+TCRαβ+-expressing double-positive cells (CD4+CD8+CD3+TCRαβ+) are shown. On average, 38% of the total viable population were DP (CD4++), of which 35% co-expressed CD3 and TCRαβ. The yields of total DP cells and CD3+TCRαβ+ DP cells per input CD34+ cell were ~23,000 and ~9,000, respectively. Shown are means with 95% confidence intervals (n = 31).



Figure 6. Frequency and Yield of CD8 SP T Cells After 49 Days of Culture

DP cells were further matured into CD8 SP T cells by culturing for an additional 7 days in StemSpan™ SFEM II with T Cell Progenitor Maturation Supplement (Catalog #09930), IL-15 (Catalog #78031) and ImmunoCult™ CD3/CD28/CD2 T Cell Activator (Catalog #10970) on coated plates. On day 49, cells were (A) analyzed by flow cytometry for the expression of CD3, TCRαβ, CD4 and CD8. The (B) frequency and yield of CD3+TCRαβ+-expressing cells and their subsets are shown. On average, 54% of the CD3+TCRαβ+ cells were DP (CD4+CD8+) and 38% were CD8 SP (CD4-CD8+). The average yield of CD8 SP T cells per input CD34+ cell was ~6,000. CD3+TCRαβ+ CD4 SP (CD4+CD8-) T cells were detected at very low frequencies (data not shown). Shown are means with 95% confidence intervals (n = 12).



Key Applications

Ex Vivo Expansion of HSPCs for Rapid/Sustained Hematopoietic Recovery Post Transplantation

Generating Target Cells for Reprogramming to Make Induced Pluripotent Stem Cells

Gene Transfer into HSPCs

SFEM
Lechman et al. (2012) Attenuation of miR-126 Activity Expands HSC In Vivo without Exhaustion. Cell Stem Cell 11(6)

SFEM II
Buechele C et al. (2015) MLL leukemia induction by genome editing of human CD34+ hematopoietic cells. Blood. Epub ahead of print
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