The Thermo Scientific Pierce Mouse Embryonic Fibroblast Isolation Kit provides isolation and culturing reagents for the optimal yield and viability of fibroblasts from mouse embryos.
The Mouse Embryonic Fibroblast Isolation Kit has been designed to obtain high viability and purity of embryonic fibroblasts using validated reagents and an optimized procedure. The kit contains a tissue-specific dissociation enzyme and media formulated for primary cell culture that promotes the isolation growth of functional fibroblasts. In addition, MEF isolated and cultured with the Pierce MEF Isolation Kit can be cryopreserved for future use with very minimal loss of viability.
- Optimized – procedure and reagents optimized for viability, yield, purity, and ease-of-use
- Yield – provides a 2-fold increase in yield compared to do-it-yourself methods and other kits
- Functional – cultured fibroblasts express the appropriate biochemical markers
- Complete kit includes Fibroblast Isolation Enzyme, Hanks' Balanced Salt Solution (HBSS), and DMEM for Primary Cell Isolation
- Generation of immortalized cell lines from transgenic mice
- Feeder layer for stem cells
- IHC and IF studies
Because it uses an optimized tissue-specific dissociation enzyme, this kit isolates fibroblasts with greater yield and cell viability than cells isolated by existing methods. Isolated MEF express fibroblast-specific protein markers, confirming their viability and unperturbed metabolic function. After isolation, the mouse fibroblasts may be immortalized by continuous passage for the establishment of unique cell lines. They may also be used as a cell culture model for a diverse range of studies such as gene regulation and stem cell research [1,2,3,4].
|Cell Yield and Viability Comparison. Cell yield and viability after cell isolation using the Thermo Scientific Pierce Mouse Embryonic Fibroblast Isolation Kit. Mouse embryonic tissue at E11-13 were minced, incubated with Pierce Mouse Embryonic Fibroblast isolation enzyme (with papain) for 30 minutes, and disrupted by pipetting with a pipette and 1000µL tip (20X) to generate single cell suspension. Cell yield and cell viability are acquired from cell suspension isolated from one mouse embryo in a total volume of 1.5mL. Cell viability is determined by trypan blue exclusion assay and total cell yield is determined using an Invitrogen™ Countess™ Automated Cell Counter.
|Representitative images of MEFs immunostained with specific protein markers. Mouse Embryonic Fibroblast were isolated and cultured using the Thermo Scientific Pierce Mouse Embryonic Fibroblast Isolation Kit. Fibroblasts were grown in 24 well plates seeded at 1 x 10^5 cells/well. At Day 7, cells were fixed with 4% paraformaldehyde, permeabilized, and stained with antibodies specific to Connexin 43 (red, upper panel, Part No. PA1-37497) or Vimentin (red, lower panel) and β-Actin (green). Nuclei (lower panels only) were visualized using Hoechst 33342 (blue)(Part No. 62249). Images were taken at 40X.
- Wang, S., et al. (2012) Inhibition of AMPKα by doxorubicin accentuates genotoxic stress and cell death in mouse embryonic fibroblasts and cardiomyocytes: role of p53 and SIRT1. J. Biol. Chem. 287(11):8001-12.
- Blackman, B.E., et al. (2009) PDE4D and PDE4B function in distinct subcellular compartments in mouse embryonic fibroblasts. J. Biol. Chem 286 (14):12590-601.
- Vierbuchen, T., et al. (2010) Direct conversion of fibroblasts to functional neurons by defined factors. Nature 463:1035-41.
- Imamura, M., et al. (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 126(4):663-76.
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