title: Fast and simple online sample preparation coupled with capillary LC-MS/MS for determination of prostaglandins in cell culture supernatants.
abstract: An online 2-D 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(2a), 6-keto-(6k)PGF(1a), and 15-Delta(12,14)-deoxy-PGJ(2) (15dPGJ(2)) in cell culture supernatants was developed and validated. Pretreatment of the cell culture supernatants included dilution and filtration, and the analysis time including all sample preparation steps was less than 50 min per sample. Peptides/proteins contained in the matrix were removed by the SCX column. LODs in the range of 0.4-2.2 ng/mL cell culture supernatant, recoveries higher than 80% and within- and between-day precisions of less than 30% RSDs were obtained. Human mesenchymal stem cells (hMSCs) were stimulated with cytokine-containing supernatants derived from activated T lymphocytes, and PG production was analyzed using the developed method. PGE(2 )was found in cultures from both untreated and stimulated hMSCs, while PGE(1) was present above the detection limit only in stimulated cells.
authors.affiliation: Institute of Immunology, Rikshospitalet-Radiumhospitalet Medical Centre, University of Oslo, N-0315 Oslo, Norway. sandrari@kjemi.uio.no
authors.forename: Sandra
authors.initials: S
authors.lastname: Rinne
authors.forename: Charlotte
authors.initials: C
authors.lastname: Ramstad Kleiveland
authors.forename: Moustapha
authors.initials: M
authors.lastname: Kassem
authors.forename: Tor
authors.initials: T
authors.lastname: Lea
authors.forename: Elsa
authors.initials: E
authors.lastname: Lundanes
authors.forename: Tyge
authors.initials: T
authors.lastname: Greibrokk
mesh_terms: D000327:Adsorption; D002478:Cells, Cultured; D002850:Chromatography, Gel / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods*; D046911:Macromolecular Substances / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D009862:Online Systems; D011108:Polymers / Q000737:chemistry; D011137:Polystyrenes / Q000737:chemistry; D011453:Prostaglandins / Q000032:analysis* / Q000302:isolation & purification; D015203:Reproducibility of Results; D012822:Silicon Dioxide / Q000737:chemistry; D012984:Software; D014867:Water / Q000737:chemistry
chemical_list: D046911:Macromolecular Substances; D011108:Polymers; D011137:Polystyrenes; D011453:Prostaglandins; D014867:Water; D012822:Silicon Dioxide