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SILAC Isotope Labeling Reagents and Kits FAQ


Answers to frequently asked questions (FAQs) about SILAC Isotope Labeling Reagents and Kits


 

What are the components of Pierce SILAC Protein Quantitation Kits?
SILAC Protein Quantitation Kits contain all reagents needed for stable isotope labeling with amino acids in cell culture (SILAC) including media, heavy and light amino acids and dialyzed FBS. We offer three classical media kits: Pierce SILAC Protein Quantitation Kit—RPMI 1640 Medium (#89982), Pierce SILAC Protein Quantitation Kit—DMEM (#89983) and Pierce SILAC Protein Quantitation Kit—DMEM F12 (#88439). Each kit includes 2 x 500mL SILAC media, 1 x 50mg L-Lysine-2HCl, 1 x 50mg 13C6 L-Lysine-2HCl, 2 x 50mg L-Arginine-HCl and dialyzed FBS.

Do you have any other heavy amino acids?
We offer additional isotopes of lysine, including 4,4,5,5 D4 L-Lysine-2HCl (#88437) and 13C6 15N2 L-Lysine-2HCl (#88209), which can be used for multiplexed experiments. 13C6 L-Arginine-HCl (#88210) and 13C6 15N2 L-Arginine-HCl (#89990) are also available to enhance peptide isotope label coverage of tryptic peptides. In addition to lysine and arginine, 13C6 L-Leucine (#88435) can also be used with our powdered SILAC DMEM (#88425) and RPMI-1640 media (#88426). Please check the product page for a current listing.

What is the recommended final concentration of the amino acids?
The amino acids supplied with the kits are used at 0.1mg/ml (50mg in 500mL media). The final concentration upon dissolving 50mg L-Lysine-2HCl or L-Arginine-HCl in 500mL medium is 0.46mM or 0.47mM, respectively. The amino acid concentrations for arginine, lysine, leucine and proline for classic media formulations are shown below. Most cell lines are not significantly affected when cultured using lower amounts of amino acids. However, some cell lines might require additional heavy or light amino acids for optimal growth and can be purchased separately. In addition, depending on the application and/or cell line, lower amounts of amino acids might be required. Some cell lines have high arginase activity, which results in the conversion of heavy arginine into heavy proline. Therefore, for best results, use the lowest concentration of arginine that supports cell growth and minimizes conversion to heavy proline. Alternatively, culture medium can be supplemented with additional proline to decrease heavy arginine conversion.

Amino acid concentrations for classic media formulations.
Media
Arginine (mM)
Lysine (mM)
Leucine (mM)
Proline (mM)
DMEM
0.4
0.8
0.8
0
DMEM F12
0.7
0.5
0.45
0.15
Ham's F12
1.0
0.2
0.1
0.3
IMDM
0.4
0.8
0.8
0.35
McCoy's 5A
0.2
0.2
0.3
0.15
MEM
0.6
0.4
0.4
0
RPMI-1640
1.1
0.2
0.4
0.2

Are the isotopes in the kit radioactive?
No. Deuterated 13C and 15N derivatives of L-rginine, L-lysine and L-leucine are stable heavy isotopes and are therefore not radioactive.

Do I need a NRC license to use this kit?
No, because the heavy isotopes used in the kits are not radioactive.

How many samples can be tested with this kit?
Each kit contains 500mL media, which can isotopically label approximately 1 x 1012 cells, depending on cell type.

If I need more of any kit component, can I purchase it separately?
We offer many of the kit components and related alternative components separately. Larger package sizes for SILAC media, dialyzed FBS and heavy and light amino acids are also available.

 

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