Human Activin Recombinant (CHO Cell)
Category: Recombinant Mouse Cytokines$380.00 – $3,700.00
Description
Accession
P08476
Source
Optimized DNA sequence encoding Human Inhibin Beta Alpha Chain was expressed in Chinese Hamster Ovary cell line.
Molecular weight
Native human Activin A is generated by the proteolytic removal of the signal peptide and propeptide, the molecule has a calculated molecular mass of approximately 13 kDa. Recombinant Activin A is a glycosylated disulfide-linked homodimeric protein consisting of two alpha chain amino acid residue subunits, and migrates as an approximately 26 kDa protein under non-reducing and reducing conditions in SDS-PAGE.
Purity
>97%, as determined by SDS-PAGE and HPLC
Biological Activity
The ED(50) was determined by the dose-dependent proliferation ofmurine MCP11 cell line was found to be in the les than 1 ng/ml.
Protein Sequence
MPLLWLRGFL LASCWIIVRS SPTPGSEGHS AAPDCPSCAL AALPKDVPNS 50 QPEMVEAVKK HILNMLHLKK RPDVTQPVPK AALLNAIRKL HVGKVGENGY 100 VEIEDDIGRR AEMNELMEQT SEIITFAESG TARKTLHFEI SKEGSDLSVV 150 ERAEVWLFLK VPKANRTRTK VTIRLFQQQK HPQGSLDTGE EAEEVGLKGE 200 RSELLLSEKV VDARKSTWHV FPVSSSIQRL LDQGKSSLDV RIACEQCQES 250 GASLVLLGKK KKKEEEGEGK KKGGGEGGAG ADEEKEQSHR PFLMLQARQS 300 EDHPHRRRRR GLECDGKVNI CCKKQFFVSF KDIGWNDWII APSGYHANYC 350 EGECPSHIAG TSGSSLSFHS TVINHYRMRG HSPFANLKSC CVPTKLRPMS 400 MLYYDDGQNI IKKDIQNMIV EECGCS 426
Endotoxin
Endotoxin content was assayed using a LAL gel clot method. Endotoxin level was found to be less than 0.1 ng/µg(1EU/µg).
Presentation
Human Activin A was lyophilized from a 0.2 μm filtered solution in 0.2M AcOH, pH 4.0.
Reconstitution
A quick spin of the vial followed by reconstitution in distilled water to a concentration not less than 0.1 mg/mL. This solution can then be diluted into other buffers
Storage
The lyophilized protein is stable for at least 2 years from date of receipt at -20° C. Upon reconstitution, this cytokine can be stored in working aliquots at 2° - 8° C for one month, or at -20° C for six months, with a carrier protein without detectable loss of activity. Avoid repeated freeze/thaw cycles.
Usage
This cytokine product is for research purposes only.It may not be used for therapeutics or diagnostic purposes.
Interactor
Interactor
Interactor
Molecular function
Molecular function
Methods
Cell Culture
- Conventional (primed) human iPSC line C1 (Whitehead Institute Center for Human Stem Cell Research, Cambridge, MA) (5 trypsinized single cells on an MEF feeder layer in hESM supplemented with ROCK inhibitor Y-27632 (10 μM).
- One or two days later, medium was switched to 5i/L/A-containing naive hESM.
- Following an initial wave of widespread cell death, dome-shaped naive colonies appeared within 10 days and could be picked or expanded polyclonally using 3–5 min treatment with Accutase (GIBCO) on an MEF feeder layer.
- Naive human pluripotent cells were derived and maintained in serum-free N2B27-based media supplemented with 5i/L/A.
- Medium (500 ml) was generated by inclusion of the following: 240 ml MEM/F12 , 240 ml (; 21103), 5 ml N2 supplement (; 17502048), 10 ml B27 supplement (; 17504044), 10 μg recombinant human LIF (made in-house), 1 mM glutamine , 1% nonessential amino acids , 0.1 mM β-mercaptoethanol , penicillin-streptomycin , 50 μg/ml BSA , and the following small molecules and cytokines: P0325901 (1 μM), IM-12 (1 μM), SB590885 (&…
hPSC Differentiation
- Single-cell suspensions of hPSCs were obtained by treating the hPSC cultures at 80% confluency with 1× TrypLE .
- Single cells were plated at an optimized density ranging from 5,000 cells/cm2 to 15,000 cells/cm2 (depending on the cell line) onto six-well plates coated with 0.5 μg/cm2 of ColIV or 0.5 μg/cm2 TenC in medium'>E8 medium supplemented with 10 μM Rho kinase inhibitor .
- After 24 hr (day 0), the medium was changed to IF9S medium (see 2+ salt , 40 μl/l monothioglycerol , 8.4 μg/l additional sodium selenite , 10 g/l polyvinyl alcohol , 1× GlutaMAX , 1× nonessential amino acids , 0.1× chemically defined lipid concentrate , 10.6 mg/l Holo-Transferrin , and 20 mg/l insulin .
- Differentiation was conducted in a hypoxic condition from day 0 to day 5, and then in a normoxic condition from day 6 to day 9 (
