Samples of urine were spiked with different concentrations of five pairs of standard d , l -phenothiazines. The Sep-Pak Vac 3 cc (200 mg sorbent, 55–105 μm particle size) C18-SPE cartridge (Waters, Milford, MA) was first conditioned with methanol (3 mL), followed by sodium phosphate buffer (3 mL, 5 mM; pH 7.0). The spiked samples were loaded onto a SPE cartridge. After that, the cartridge was washed with 1 mL method:water (50:50 v/v) and 2 mL deionized water. Final eluted was carried out with 3 mL methanol (100%). The eluted samples were evaporated in a centrifugal vaporizer (EYELA, CVE-200D). The dried samples were dispersed by adding a solution containing 15 μL methanol and 15 μL deionized water. Finally, the purified sample was analyzed by CE.
Sep pak vac 3 cc
Manufactured by Waters Corporation
The Sep-Pak Vac 3 cc is a solid-phase extraction (SPE) cartridge designed for sample preparation and analyte purification. It features a 3 cc volume and is intended for use in a variety of laboratory applications.
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2 protocols using sep pak vac 3 cc
1
Quantitative Analysis of Phenothiazines in Urine
2
iTRAQ 8-plex Labeling Protocol
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The iTRAQ Reagents 8-plex kit (Applied Biosystems, CA) was used according to the manufacturer’s instructions, with slight modifications. For labelling, each vial of the iTRAQ Reagents 8-plex kit was allowed to come to room temperature. Then, each tube was spun to bring the solution to the bottom of the vial. A 50-μl volume of isopropanol was added to each room-temperature vial. Each vial was vortexed and then spun. The contents of each freshly prepared iTRAQ Reagents 8-plex vial were transferred to an individual sample tube, and each tube was vortexed and then spun. The pH was tested, and up to 5 μl of dissolution buffer was added to adjust the pH to between 7.5 and 8.5. The tubes were incubated at 25°C for 4 h. The contents of each iTRAQ Reagent8-plex-labelled sample tube were transferred into one tube, and the tube was vortexed and then spun. The resulting peptides were lyophilized to dryness, desalted with C18 cartridges (Sep-Pak Vac 3cc, Waters) and concentrated by vacuum centrifugation before the next step. The correspondence between the samples and the labels is as follows: 6:00 –tag 113, 9:00 –tag 114, 11:00 –tag 115, 12:00 –tag 116, 13:00 –tag 117, 14:00 –tag 118, 16:00 –tag 119 and 18:00 –tag 121.
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