Highly automated nano-LC/MS-based approach for thousand cell-scale quantification of side chain-hydroxylated oxysterols[S]
| Autor: | Hanne Roberg-Larsen, Kaja Lund, Tore Vehus, Nina Solberg, Caroline Vesterdal, Dorna Misaghian, Petter Angell Olsen, Stefan Krauss, Steven Ray Wilson, Elsa Lundanes |
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| Jazyk: | angličtina |
| Rok vydání: | 2014 |
| Předmět: | |
| Zdroj: | Journal of Lipid Research, Vol 55, Iss 7, Pp 1531-1536 (2014) |
| Druh dokumentu: | article |
| ISSN: | 0022-2275 91617898 |
| DOI: | 10.1194/jlr.D048801 |
| Popis: | Iso-octyl chain-hydroxylated oxysterols were determined in attomoles per 10,000 cells concentrations in 10,000–80,000 cultured pancreatic adenocarcinoma cells, using a sensitive, highly automated nano-LC-ESI-MS-based method. Identified oxysterols included 24S hydroxycholesterol (24S-OHC), 25 hydroxycholesterol (25-OHC), and 27 hydroxycholesterol (27-OHC), while 20S hydroxycholesterol and 22S hydroxycholesterol were not detected. Lower mass limit of quantification was 23 fg (65 amol) for 25-OHC and 27-OHC (100 times lower than our previous method) and 54 fg (135 amol) for 24S-OHC, after derivatization into Girard T hydrazones and online sample cleanup using simplified and robust automatic filtration and filter back flushing solid phase extraction LC/MS/MS. The instrument configuration was easily installed using a commercial nano-LC/MS system. Recoveries in spiked sample were 96, 97, and 77% for 24S-OHC, 25-OHC, and 27-OHC, with within- and between-day repeatabilities of 1–21#x0025; and 2–20#x0025; relative SD, respectively. The study demonstrates the potential of nano-LC in lipidomics/sterolomics. |
| Databáze: | Directory of Open Access Journals |
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