Proteomics

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A multi factor comparison of seven reversed-phase C18 separation media for proteomic applications


ABSTRACT: This study examines the differences in separation selectivity and carryover characteristics for seven popular C18 columns used in bottom-up proteomics. Identical linear water:acetonitrile gradients (0.48 %ACN per minute) were applied to all separations at 300 nL/min flow rate. Both buffers A (water) and B (80:20 acetonitrile:water) contained 0.1 % formic acid. Columns 1-6 were packed using a slurry method into 75 mm x 50 cm PicoFrit capillaries with 10 mm pulled emitters. Columns used: (1) Reprosil Pur C18 AQ 3 mm 120 A ; (2) Reprosil Pur ODS-3 3 mm 120 A ; (3) Luna C18(2) 3 mm 100 A ; (4) XBridge BEH C18 2.5 mm 130 A ; (5) XSelect CSH 2.5 mm 130 A ; (6) ProntoSIL-C18 AQ 3 mm 200 A (also known as Magic C18 AQ) ; (7) PepMap RSLC C18, 2 mm, 100 A, 75 mm x 50 cm . 1D and 2D LC-M/MS analyses have been performed for all seven columns. 1D analyses each used ~500 ng of whole cell Jurkat tryptic digests in triplicates; the 2D analysis each spanned 7 concatenated fractions from the first dimension (high pH RP HPLC) of whole cell Jurkat tryptic digest, again with all fractions containing ~500 ng of peptides. All samples were spiked with peptide retention standard (P1-P6) for the accurate alignment of the retention data in HI (%ACN) values.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Oleg Krokhin  

PROVIDER: MSV000096640 | MassIVE | Wed Dec 11 10:28:00 GMT 2024

REPOSITORIES: MassIVE

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