SILAC kits and reagents for human mesenchymal stem cell proteomic analysis.

The Thermo Scientific Pierce Human Mesenchymal Stem Cell SILAC Kit, Media and Reagents are specifically designed for analysis of protein expression by mass spectrometry using a method called stable isotope labeling with amino acids in cell culture (SILAC). The kit includes a complete set of specialized media, supplements and heavy and light isotopes of lysine for performing a complete experiment. In addition, individual components are offered, including five different expansion and differentiation media, dialized FBS supplement and alternative lysine and arginine isotope pairs.

What is SILAC?

Stable isotope labeling with amino acids in cell culture (SILAC) is a powerful method to identify and quantify relative differential changes in complex protein samples. The SILAC Method uses in vivo metabolic incorporation of "heavy" ¹³C- or ¹⁵N-labeled amino acids into proteins followed by mass spectrometry (MS) analysis for accelerated comprehensive identification, characterization and quantitation of proteins.

... Learn more about the SILAC method.

Highlights of SILAC Products for Human Mesenchymal Stem Cells:

• Optimized, specialized media – the Universal Expansion Media is specifically designed and qualified for human mesenchymal stem cell propagation and SILAC protein expression profiling with mass spectrometry; alternatively, choose one of several compatible differentiation media:
  • Adipogenic Differentiation Media
  • Chondrogenic Differentiation Media
  • Neural Differentiation Media
  • Osteogenic Differentiation Media
• High-quality supplements – the Stem Cell Screened Dialyzed FBS is tested to ensure that it does not promote stem cell differentiation; the Phenol Red-Free MEM Media for SILAC can be used with estrogen responsive cells such as the breast adenocarcinoma cell line MCF7, without inducing an artificial estrogen response
• Flexible and compatible – media and the SILAC procedure are compatible with all standard mass spectrometry instrumentation
• Multiplex capabilities – several alternative isotopes of arginine and lysine are available that allow for analysis of multiple growth conditions in each experiment. (For additional individual SILAC reagents and amino acid isotopes see related SILAC Product page).
• Thousands of proteins – in vivo labeling with arginine and lysine isotopes using endogenous translational machinery enables expression analysis of the entire proteome; investigate any protein or group of proteins you choose

SILAC Applications:

• Quantitative analysis of relative changes in protein abundance from different cell treatments
• Quantitative analysis of proteins for which there are no antibodies available
• Protein expression profiling of normal vs. disease cells
• Identification and quantification of hundreds to thousands of proteins in a single experiment

Differentiation of human adipose derived mesenchymal stem cells (hAMSC) for SILAC workflows. A. Undifferentiated hAMSC were expanded in universal expansion media for SILAC supplemented with "light"or "heavy" arginine and lysine. Cell populations grown in "light" expansion media were differentiated using Adipogenic Differentiation Media (#88202) (B-D). Differentiation was visualized with the following IHC markers: B. oil red O staining of adipocytes C. beta III Tubulin staining of neurons D. phallodin staining of osteocytes.

SILAC Poster Presentations

• Hughes, D.E., Hong, S.J., Rosenblatt, M., Krastins, B., Sarracino, D., Lopez, L., Kamath, A., Ghosh, R. and Rogers, J. (2008). Characterization of human mesenchymal stem cell neuronal differentiation by SILAC and high content screening analysis. Mol. Biol. Cell 19 (suppl), abstract # 2862/B578.

The following posters were presented at the American Society for Mass Spectrometry, June 2007:

• Quantitation of Chemical and Genetic Alteration of the Proteome using Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC)
• Protein Quantitation of p53-Regulated Proteins using Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC)
• Using Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC) to Quantitate Exportin/CRM1-dependent Protein Transport in Nuclear Extracts

SILAC Literature References

1. Amanchy, R. et al. (2005). Science STKE 267: 1-20
2. Blagoev, B., et al. (2004). Nat Biotechnol 22(9): 1139-1145.
3. Everley, P. et al. as presented at The American Society for Biocehmistry and Molecular Biology. MCP Papers in Press. Published on July 11, 2007 as Manuscript M700057-MCP200.
4. Kratchmarova, I., et al. (2005). Science 308(5727): 1472-1477.
5. Mann, M. (2006). Nat Rev Mol Cell Biol 7(12): 952-958.
6. Ong, S. E., et al. (2002). Mol Cell Proteomics 1(5): 376-386.
7. Selbach, M. and Mann, M. (2006). Nat. Methods 3(12): 981-3.

Related Products and Links
Individual SILAC Reagents – including universal supplements and additional amino acid isotopes
Standard SILAC Kits – including DMEM and RPMI-160 media for SILAC
HyClone AdvanceSTEM Human Somatic Stem Cells – compatible validated cells
Open Biosystems viPS Vector Kit – for reprogramming somatic cells to generate induced pluripotent stem cells
AQUA and Heavy Peptides – custom peptide isotopes for mass spectrometry
Silver Stain for Mass Spectrometry – exceptional sensitivity and recoverability for MS
In-Gel Trypsin Digest Kit – optimized kit for protein MS sample preparation
Prepare SILAC peptides using the In-Gel Tryptic Digestion Kit (Tech Tip #60)

The purchase of this product conveys a non-transferable license to the Purchaser to use this product in methods protected under U.S Patent 6,653,076 (owned by University of Washington) for research purposes only.