Iodomethane d3

Silver oxide, anhydrous sodium sulfate, iodomethane, iodomethane-d₃, l-phenylalanine methyl ester hydrochloride, mass spectrometric grade formic acid (~98%), and HPLC grade trifluoroacetic acid (99.0%) were obtained from Sigma-Aldrich; HPLC grade methanol, acetonitrile, ethyl acetate, and water from Fisher Scientific; Deuterium oxide (99.8%) and HPLC grade dichloromethane from Acros; l-alanyl-l-proline, N-(tert-butoxycarbonyl)-d-alanine, l-proline tert-butyl ester hydrochloride, d-proline tert-butyl ester hydrochloride, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) hydrochloride from Tokyo Chemical Industry. A Fisher Scientific vortex mixer was used for mixing and a Sorvall Legend Micro 21R microcentrifuge used for centrifugation of 1.5 mL Eppendorf tubes. A Corning Laboratory stirrer was used for mixing chemical reactions. Human plasma (Na₂-EDTA) was obtained from Bioreclamation and stored at −80 °C. An Argonaut SPE DRY™ 96 DUAL evaporator was used for solvent evaporation.

HPLC/analytical grade reagents were used throughout. 1,1-carbonydiimidiazole, triethylamine, iodomethane, iodomethane-d3, iodoethane, iodoethane-d5, iodopropane, formic acid, citric acid, methanol, disodium hydrogen phosphate, chloroform, ammonium formate, acetonitrile, 1-butanol, and analytical standards for saturated and unsaturated fatty acids were purchased from Sigma-Aldrich. All lipid extractions were performed in 2 mL polypropylene LoBind safe-lock tubes (Eppendorf).
Cell culture reagents were purchased from Life Technologies. Mouse anti-STXBP1 antibody was purchased from BD Biosciences. pCMV-STXBP1-emGFP, pCMV-STXBP1, NPY-hPLAP, and NPY-mCherry were prepared as previously described (Arunachalam et al, 2008 (link); Martin et al, 2013 (link); Tomatis et al, 2013 (link)). STXBP1^Δ317-333 and STXBP1^F115E mutants were made using the quick-change lightning site-directed mutagenesis kit (Strategene, USA) and the mutational primer
5′-GACTTTTCCTCTAGCAAGAGGATGATGCCCCAGTACCAGAAGGAGC-3′, as previously described (Martin et al, 2013 (link)). All constructs were sequenced at The Australian Genome Research Facility, located at The University of Queensland.

Saturated and unsaturated FFAs were obtained from Sigma-Aldrich. Phospholipid standards (PC 17:0/17:0, PE 17:0/17:0, PS 17:0/17:0, PA 17:0/17:0, PG 17:0/17:0, PC 12:0/13:0, PE 17:0/14:1, PS 17:0/14:1, PA 12:0/13:0 and PG 12:0/13:0) were purchased from Avanti Polar Lipids. 1,1-carbonydiimidiazole, 1,4 dioxane, triethylamine, iodomethane, iodomethane-d₃, iodoethane, formic acid, citric acid, disodium hydrogen phosphate ammonium formate, acetonitrile, 1-butanol, methanol and chloroform were purchased from Sigma Aldrich. All reagents were analytical grade or equivalent. All lipid extractions were performed in 2 mL polypropylene LoBind safe-lock tubes (Eppendorf).