Twenty microliters of perfusate was mixed with 10 μl of borate buffer 0.5 mol/l and 5 μl of fresh solution [4 mg ophtalaldehyde (OPA) and 2 μl of β-mercaptoethanol in 1 ml of methanol] for derivatization. The mixture was agitated for 30 s and injected 1 min later in the HPLC column (LiChroCART 250-4 Purospher STAR RP-18 endcapped, particle size 5 μm; Merck KGaA, Darmstadt, Germany).The mobile phase consisted of phosphate buffer (0.1 mol/l KH2PO4) with an acetonitrile gradient from 25 to 50% at a flux of 1 ml per minute. Acetonitrile and phosphate buffer were mixed by the pump (Elite labChrom L-2130; Hitachi, Chiyoda, Tokyo, Japan) in an automatized manner. The fluorescent derivatives were analysed using the integrated fluorometric detector (Elite labChrom L-2480; Hitachi). Excitation/emission wavelengths used were 330/450 nm. The calibration curve was constructed using histamine and serotonin (from Sigma-Aldrich Co.) standards dissolved in artificial CSF in a concentration range from 0.06 to 0.25 pmol/20 μl, which yielded a linear correlation between neurotransmitter concentration and peak area.
Lichrocart 250 4 purospher star rp 18 endcapped
Manufactured by Merck Group
Sourced in Germany
The LiChroCART 250-4 Purospher STAR RP-18 endcapped is a type of liquid chromatography column. It is designed for the separation and analysis of a variety of organic compounds. The column is packed with a silica-based stationary phase that is chemically modified with octadecyl (C18) functional groups. This stationary phase is suitable for the reversed-phase liquid chromatography technique.
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Lab products found in correlation
2 protocols using lichrocart 250 4 purospher star rp 18 endcapped
1
Quantification of Biogenic Amines in Perfusate
2
Quantification of Histamine and Serotonin
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Ten microliters of borate buffer 0.5 M and 5 μl of the fresh solution [4 mg ophtalaldehyde (OPA) and 2 μl of β-mercaptoethanol in 1 ml of methanol] were added to 20 μl of the microdialysis sample for derivatization. The mixture was agitated for 30 s and injected after 1 min in the HPLC column (LiChroCART 250-4 Purospher STAR RP-18 endcapped, particle size 5 μm; Merck KGaA, Darmstadt, Germany). The mobile phase contained phosphate buffer (0.1 mol/L KH2PO4) and an acetonitrile gradient from 25 to 50% and was set at a flux of 1 ml per minute. Acetonitrile and phosphate buffer were automatically mixed by the pump (Elite labChrom L-2130; Hitachi, Chiyoda, Tokyo, Japan). The resultant fluorescent derivatives were analyzed using a fluorometric detector (Elite labChrom L-2480; Hitachi). We used excitation/emission wavelengths of 330/450 nm, respectively. Histamine and serotonin (from Sigma-Aldrich, Co.) standards dissolved in ACSF in a concentration range from 0.06 to 0.25 pmol/20 μl were used to construct calibration curves that produced a linear correlation between neurotransmitter concentration and peak area.
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