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Detecting pM concentrations of prostaglandins in cell culture supernatants by capillary SCX‐LC‐MS/MS
Author(s) -
Rinne Dahl Sandra,
Ramstad Kleiveland Charlotte,
Kassem Moustapha,
Lea Tor,
Lundanes Elsa,
Greibrokk Tyge
Publication year - 2008
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.200800184
Subject(s) - chromatography , chemistry , mass spectrometry
A highly sensitive, improved online strong cation exchange (SCX) – RP capillary liquid chromatographic (cLC) method with IT mass spectrometric (IT‐MS/MS) detection for the simultaneous determination of prostaglandin (PG)A 1 , PGD 2 , PGE 1 , PGE 2 , PGF 2α , 8‐iso‐(8i)PGF 2α , 6‐keto‐(6k)PGF 1α , and 15‐Δ 12, 14 ‐deoxy‐PGJ 2 (15dPGJ 2 ) in cell culture supernatants was developed and validated. Pretreatment of the cell culture supernatants included only dilution and filtration, and the analysis time including all sample preparation steps was 60 min per sample. Peptides/proteins contained in the matrix were removed by the SCX column. LODs in the range of 8–44 pg/mL (25–120 pM) cell culture supernatant were obtained. Excellent linearity ( R 2 > 0.99) and satisfactory recoveries and within‐ and between‐day precisions were obtained. Human mesenchymal stem cells (hMSCs) were stimulated with tumor necrosis factor α (TNFα) or TNFα/IL‐17, and PG production was analyzed using the developed method. The four PGs, 6kPGF 1a , PGF 1a , PGE 2 , and PGE 1 were detected both in nonstimulated and stimulated cells. The amount of PG produced by the cell increased when the cell was stimulated.