Apr 30, 2026

SILAC DMEM 1 recipe

This  protocol  is a draft, published without a DOI.
  • 1Discovery Research Platform for Hidden Cell Biology
Victoria Munro
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Protocol Citation: Georg Kustatscher 2026. SILAC DMEM 1 recipe. protocols.io https://dx.doi.org/
License: This is an open access  protocol  distributed under the terms of the  Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: In development
We are still developing and optimizing this protocol
Created: June 21, 2024
Last Modified: April 30, 2026
Protocol  Integer ID: 102219
Keywords: standard dmem for silac media, silac dmem, thaw 500ml bottle, silac media, ml in ultrapure water, precipitates amino acid, recipe media recipe, standard dmem, cell culture media, abundant peptide, protein gradient, amino acid, freeze thaw cycle, lysine, 50ml aliquot, ultrapure water
Abstract
Media recipe for our standard DMEM for SILAC media.

Notes:
  • Van originally used 50mg/500ml for all RKP but there was a very small amount of R10->P6 conversion visible in MS1 for abundant peptides
  • Dialysed FBS: Label with dialysed FBS batch so that L and H media with the same dFBS can be used in experiment. Freeze thaw cycles should be limited, so thaw 500ml bottle and make 50ml aliquots for adding to cell culture media and store at -20°C. Thaw at 4°C or at 37°C, ensuring to mix throughout thawing to prevent salt/protein gradients which increase precipitates
  • Amino acids: Make up arginine, lysine and proline to 0.1g/ml in UltraPure water (Invitrogen; 10977035), filter-sterilise using 0.2um filters and aliquot
SILAC MEDIA RECIPE
500ml DMEM for SILAC (Thermo A33822)
  • Deficient in arginine and lysine 
50ml dialysed FBS (Gibco, 26400044)
  • Spin 3202g 3 min before adding to remove precipitate
5ml 100X Penicillin-Streptomycin (Gibco 11548876)
5ml 100X Glutamax (Thermo 35050061)
Lysine (Heavy Lysine.HCl; Sigma 608041 or Light Lysine.HCl Sigma, L8662-100G)
  • 500ul heavy from 0.1g/ml stock (50mg/500ml; 100mg/L; 525uM)
  • 479.2ul light from 0.1g/ml stock (47.94mg/500ml; 95.834mg/L; 525uM)
Arginine (Heavy Arginine.HCl: Sigma, 608033 or Light Arginine: Sigma, A8094-100G)
  • 250ul heavy from 0.1g/ml stock (25mg/500ml; 50mg/L; 227uM)
  • 197.4ul light from 0.1g/ml stock (19.74mg/500ml; 39.485mg/L; 227uM)
Proline (Sigma, P5607-25G)
  • 2ml from 0.1g/ml stock (200mg/500ml; 400mg/L; 3.47mM)