This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Analysis of mannose-6-phosphate by High pH Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD) Each of the samples was dissolved with 30 ?L of nanopure H2O and injected at 10 ?L. However, sample, Fc4 FK, was re-injected at 15 ?L because no peak for mannose-6-phosphate was observed in the first injection (10 ?L). Four concentrations of mannose-6-phosphate standard (80 pmoles/10 ?L, 160 pmoles/10 ?L, 320 pmoles/10 ?L and 640 pmoles/10 ?L) were prepared by serial dilutions. A water-blank was injected in addition to the prepared samples and a calibration curve was established based on the area units generated from each concentration. The samples and standards were analyzed by HPAEC using a Dionex DX500 system equipped with a GP40 gradient pump, an ED40 electrochemical detector, Dionex CarboPac PA20 (3 x 150 mm) analytical column and a Thermo-Separation AS3500 autosampler containing a stainless steel needle. The autosampler was set to deliver an injection volume of 10 ?L (except for the re-injections of Fc4 FK) and injection was made every 35 minutes. The eluents used in the gradient program were 100 mM NaOH and 1 M sodium acetate in 100 mM NaOH.
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