S-Trap eliminates Pluronic F68 and cell culture media polymers for effective proteomic analysis of mammalian cell bioreactor supernatants

ABSTRACT: Proteomic analysis of bioreactor supernatants can inform on cellular metabolic status, viability, and productivity, as well as product quality, which can in turn help optimize bioreactor operation. Incubating mammalian cells in bioreactors requires the addition of polymeric surfactants such as Pluronic F68, which reduce the sheer stress caused by agitation. However, these surfactants are incompatible with mass spectrometry proteomics and must be eliminated during sample preparation. Here, we compared four different sample preparation methods to eliminate Pluronic F68 from filtered bioreactor supernatant samples: organic solvent precipitation; filter-assisted sample preparation (FASP); S-Trap; and single-pot, solid-phase, sample preparation (SP3). We found that SP3 and S-Trap substantially reduced or eliminated the polymer(s), but S-Trap provided the most robust clean-up and highest quality data. Additionally, we observed that SP3 sample preparation of our samples and in other published datasets was associated with partial alkylation of cysteines, which could impact the confidence and robustness of protein identification and quantification. Finally, we observed that several commercial mammalian cell culture media and media supplements also contained polymers with a similar mass spectrometry profile to Pluronic F68, and we suggest that proteomic analyses in these media will also benefit from the use of S-Trap sample preparation.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Homo Sapiens (human) Saccharomyces Cerevisiae (baker's Yeast)

TISSUE(S): Cell Suspension Culture

SUBMITTER: Lucia Zacchi  

LAB HEAD: Benjamin Schulz

PROVIDER: PXD017214 | Pride | 2020-06-30

REPOSITORIES: Pride