Stemfactor™ BMP-4, Human Recombinant

Bone Morphogenetic Protein 4 (BMP-4) is a polypeptide belonging to the TGFβ protein super-family. BMP-4 is involved in bone and cartilage development; more specifically, in tooth and limb development fracture repair¹. In human embryonic development, BMP-4 is a critical signaling molecule required for the early differentiation of the embryo and establishment of a dorsal-ventral axis^2,3. BMP-4 plays an important role in the differentiation of overlying ectodermal tissue. Inhibition of the BMP-4 signal causes the ectoderm to differentiate into the neural plate. In cultured stem cells, BMP-4 plays a distinct role in mouse and human embryonic stem (ES) cells. BMP-4 supports LIF as a positive factor for mouse ES cell self-renewal⁴. In contrast, BMP-4 induces extra-embryonic trophoblast differentiation in human pluripotent stem cells⁵. Stemfactor BMP-4 is a recombinant protein expressed and purified from human 293 cells as a glycosylated homodimer with a molecular mass of 34 kDa.

Stemgent and the Stemfactor brand name are trademarks of REPROCELL Inc., Japan.

Product Name: Stemfactor BMP-4, Human Recombinant

Catalog Number: 03-0007

Size: 10 µg

Purity: Greater than 95% by SDS-PAGE analysis.

Formulation: Lyophilized from sterile-filtered 1 M NaCl, 50 mM NaOAc, pH 4.5.

Reconstitution:

• 2019 (Lot J1809-04) or earlier: Centrifuge briefly and then reconstitute BMP-4 in 4 mM HCl to yield a stock solution of no less than 0.1 mg/mL. Avoid freeze-thaw cycles as they can result in loss of activity.
• 2020 (Lot J1912-05) or later: Centrifuge vial briefly before opening. Suspend BMP-4 in sterile water by gently pipetting water down the sides of the vial. DO NOT VORTEX. Allow several minutes for complete suspension. For prolonged storage, dilute to a working dilution in 0.1% BSA in water, divide into single-use aliquots, and store at -80 °C. Avoid repeated freeze-thaws.

Storage and Stability: Stemfactor BMP-4 is shipped at room temperature. Lyophilized BMP-4 is stable for up to 6 months from date of receipt when stored at −20 °C to −80 °C.

• 2019 (Lot J1809-04) or earlier: Reconstituted BMP-4, at concentrations greater than or equal to 0.1 mg/mL, is stable for up to 3 months when stored at −20 °C and up to 6 months when stored at −80 °C.
• 2020 (Lot J1912-05) or later: Reconstituted solutions in water are stable for 1 month at 4 °C. Working dilutions in BSA solution are stable for 3 months at -20 °C to -80 °C.

Sterility: Tested to be negative for Mycoplasma sp. by PCR and microbial contamination by a sterility test.

Source: Stemfactor BMP-4 was expressed in and purified from human 293 cells.

Amino Acid Sequence: SPKHHSQRAR KKNKNCRRHS LYVDFSDVGW NDWIVAPPGY QAFYCHGDCP FPLADHLNST NHAIVQTLVN SVNSSIPKAC CVPTELSAIS MLYLDEYDKV VLKNYQEMVV EGCGCR

Uniprot Accession Number: P12644, residues 293-408

Endotoxin Level: Less than 1.0 EU/µg of BMP-4 as determined by the LAL method.

Biologic Activity: The ED50 is less than 30 ng/mL as determined by its ability to induce alkaline phosphatase production by mouse chondrogenic ATDC-5 cells.

Intended use: This product is intended for research use only, and is not for therapeutic or diagnostic use in humans.

Specification Sheets:

• 03-0007 Specifications Sheet

Safety Data Sheets:

• 03-0007 Safety Data Sheet

1. Miljkovic, N.D., Cooper, G.M., and Marra, K.G. Chondrogenesis, bone morphogenetic protein-4 and mesenchymal stem cells. Osteoarthritis Cartilage 16: 1121-1130 (2008).
2. Chen, D., Zhao, M., and Mundy, G.R. Bone morphogenetic proteins. Growth Factors 22: 233-241 (2004).
3. Sadlon, T.J., Lewis, I.D., and D'Andrea, R.J. BMP4: its role in development of the hematopoietic system and potential as a hematopoietic growth factor. Stem Cells 22: 457-474 (2004).
4. Ying, Q.L., Nichols, J., Chambers, I., and Smith, A. BMP induction of Id proteins suppresses differentiation and sustains embryonic stem cell self-renewal in collaboration with STAT3. Cell 115: 281-292 (2003).
5. Xu, R.H., Chen, X., Li, D.S., Li, R., Addicks, G.C., Glennon, C., Zwaka, T.P., and Thompson, J.A. BMP4 initiates human embryonic stem cell differentiation to trophoblasts. Nat Biotechnol 20: 1261-1264 (2002).

Additional Publications

• Mao X; An Q; Xi H; Yang X-J; Zhang X; Yuan S; Wang J; Hu T; Liu Q; Fan G. Single-Cell RNA Sequencing of hESC-Derived 3D Retinal Organoids Reveals Novel Genes Regulating RPC Commitment in Early Human Retinogenesis. Stem Cell Rep in press :e2609 (2019).
• Schaefer SA; Higashi A; Loomis B; Schrepfer T; Wan G; Corfas G; Dressler GR; Duncan RK. From Otic Induction to Hair Cell Production: Pax2^EGFP Cell Line Illuminates Key Stages of Development in Mouse Inner Ear Organoid Model. Stem Cells Development 27:237-251 (2018).
• Koehler KR; Ine J; Longworth-Mills E; Liu X-P; Lee J; Holt JR; Hashino E. Generation of inner ear organoids with functional hair cells from pluripotent stem cells. Nat Biotechnol 35:583-589 (2017).
• Noack C; Haupt LP; Zimmerman W-H; Streckfuss-Bomeke; Zelarayan LC. Generation of a KLF15 homozygous knockout human embryonic stem cell line using paired CRISPR/Cas9n, and human cardiomyocytes derivation. Stem Cell Research 23:127-131 (2017).
• Saxena S; Ronn RE; Guibentif C; Moragheba R; Woods N-B. Cyclic AMP signaling through Epac axis modulates human hemogenic endothelium and enhances hematopoeitic cell generation. Stem Cell Rep doi:10.1016/j.stemcr.2016.03.006 (2016).
• Qian X; Kim JK; Tong W; Villa-Diaz LG; Krebsbach PH. DPPA5 supports pluripotency and reprogramming by regulating NANOG turnover. Stem Cells 34:588-600 (2016).
• Qian X. "Defining the mechanism by which synthetic polymer surfaces support human pluripotent stem cell self-renewal." PhD Thesis, University of Michigan (2015).
• Zhu S; Wang H; Deng S. Reprogramming fibroblasts toward cardiomyocytes, neural stem cells and hepatocytes by cell activation and signaling-directed lineage conversion. Nature Protocols 10, 959-973 (2015).
• Villa-Diaz LG; Kim JK; Lahann J; Kresbach PH. Derivation and long-term culture of transgene-free human induced pluripotent stem cells on synthetic substrates. Stem Cells Trans Med 3:1410 (2014).
• Chiang P-M; Wong PC. Differentiation of an embryonic stem cell to hemogenic endothelium by defined factors: Essential role of bone morphogenetic protein 4. Development 138: 2833-2843 (2011).
• Wang, B., Miyagoe-Suzuki, Y., Yada, E., Ito, N., Nishiyama, T., Nakamura, M., Ono, Y., Motohashi, N., Segawa, M., Masuda, S., Takeda, S. Reprogramming Efficiency and Quality of Induced Pluripotent Stem Cells (iPSCs) Generated from Muscle-derived Fibroblasts of MDX Mice at Different Ages. PLOS Currents Muscular Dystrophy. 2011 Oct 27 . Edition 1. doi: 10.1371/currents.RRN1274 .
• Yanes O; Clark J; Wong DM; Patti GJ; Sanchez-Ruiz A; Benton HP; Trauger SA; Desponts C; Ding S; Siuzdak G. Metabolic oxidation regulates embryonic stem cell differentiation. Nature Chemical Biology 6:411 (2010).