YILIMART - Human SHH, premium gr. (25 μg)

Recombinant Human SHH (Sonic Hedgehog; or C24II) can promote growth and differentiation of embryonic stem cells, adult mesenchymal stem cells, and cells within the neuronal lineage. The morphogenetic cytokine is crucial for embryonic development, as it is involved in the patterning of the central nervous system, somites, and limbs. Recombinant Human SHH is ideal for use in cell culture, differentiation studies, and functional assays.

Applications

Human SHH (C24II) can be used for a variety of applications including:

Expansion or differentiation of various cell types, mainly human and mouse embryonic stem cells, but also adult stem cells and mesenchymal cells.

Background information

Sonic hedgehog (SHH) is involved in a wide variety of embryonic developmental events, including neurogenesis, limb development, hematopoiesis, and gut formation. Besides their role as morphogens, hedgehog family members have also been shown to act as mitogens, cell survival factors, and axon guidance factors and to influence tissue regeneration within the adult. SHH is synthesized as a precursor, which is autocatalytically cleaved into a 19 kDa N-terminal domain (SHH-N) and a 25 kDa C-terminal domain (SHH-C). SHH-N is responsible for the signaling activity of hedgehog proteins, while SHH-C acts as a cholesterol transferase and covalently attaches a cholesterol molecule to SHH-N. The lipid-modified SHH-N displays enhanced activity. Human SHH (C24II) corresponds to the mature SHH-N domain and carries a Cys to Ile-Ile substitution resulting in a lipophilic moiety and therefore an increased activity.

Quality description

Research-grade

cytokines are suitable for a wide variety of cell culture applications. They are sterile-filtered prior to lyophilization. Generally, endotoxin levels are <0.1 ng/μg (<1 EU/μg), and purities are >95%. The biological activity is tested in appropriate bioassays.

Premium-grade

cytokines offer the convenience of high and well-defined biological activities and allow exact unit dosing for demanding applications. The biological activity is determined after lyophilization and reconstitution, and normalized to WHO/NIBSC standards whenever available. In general, endotoxin levels are <0.01 ng/μg (<0.1 EU/μg), and purities are >97%. Lot-specific certificates of analysis are available on request (macstec@miltenyibiotec.de).

Biological activity

Alkaline phosphatase production by mouse C3H10T1/2 fibroblasts
premium grade: ≥ 2×

10
³
U/mg
research grade: ≥ 1×

10
³
U/mg

Figure 1

Neural precursors, derived from mouse embryonic stem cells, were expanded in MACS Neuro Medium containing N2-supplements, laminin, Human FGF-2 (10 ng/mL), Human SHH (C24II) (200 ng/mL), and FGF-8b (100 ng/mL). Subsequently cells were grown in MACS Neuro Medium complemented by MACS NeuroBrew-21 and ascorbic acid (200 µM) for final differentiation into neurons. Cells were analyzed by immunocytochemistry for TuJ1, TH, and GAD67 expression.

TH (tyrosine hydroxylase) ⁺ cells	TuJ1 (neuron-specific β-III tubulin) ⁺ cells	GAD67 (glutamate decarboxylase) ⁺ cells

Figure 2
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Figure 2

Human SHH (C24II) activity assay.
The biological activity of Human SHH (C24II) is determined by alkaline phosphatase assay using mouse C3H10T1/2 fibroblasts.

Human SHH (C24II) activity assay.
The biological activity of Human SHH (C24II) is determined by alkaline phosphatase assay using mouse C3H10T1/2 fibroblasts.
Figure 3
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Figure 3

Mass spectrometry analysis (ESI-MS) of Human SHH (C24II), premium grade. The peak corresponds to the calculated molecular mass of 19683 Da.

Mass spectrometry analysis (ESI-MS) of Human SHH (C24II), premium grade. The peak corresponds to the calculated molecular mass of 19683 Da.
Figure 4
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Figure 4

SDS-PAGE of Human SHH (C24II), premium grade
under reduced (R) and non-reduced (NR) conditions.

SDS-PAGE of Human SHH (C24II), premium grade
under reduced (R) and non-reduced (NR) conditions.

Selected references
1. Nakamura, T. et al. (1997) Induction of osteogenic differentiation by hedgehog proteins. Biochem. Biophys. Res. Commun. 237: 465-469
Certificates

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Human SHH, premium gr. (25 μg)

For research use only

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Applications

Background information

Quality description

Biological activity

Figure 1

Figure 2

Figure 2

Figure 3

Figure 3

Figure 4

Figure 4

Selected references

Certificates

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