ANYL 190 |
| Neuropeptides play a significant role in many physiological processes and are therefore very important biomarkers with respect to neurological disorders. Quantitation of neuropeptides is very difficult due to their low concentrations (<50pg/mL), poor stability (half lives for some of 1-3 minutes in blood) and interference from the biological media. In the past nano-LC/MS/MS techniques have been tried with limited success, the main drawback being the lack of a stable MS signal resulting in RSDs of greater then 20%. When comparing recently developed emitter/column combinations and various nanospray sources (all with optimization of flow rates and applied voltage and continuous rinsing of the injector to emitter path) we observed lower RSDs as well as sufficiently low detection limits suggesting that this technique can be used for quantitation of low level neuropeptides. To take advantage of the new technology and to insure success, correct preparation of the samples is also crucial. In our first attempt at isolation of neuropeptides we denatured CSF and Plasma samples with methanol at 4oC for several hours followed by ultra filtration of the resulting mix. The resulting filtrate was dried down and taken up in the appropriate buffer and analyzed by nano-LC/MS/MS. Quantifiable levels of three different neuropeptides were observed at concentrations between 25 to 75 pg/mL. Work is ongoing to further demonstrate the feasibility of quantifying neuropeptides using nano-LC/MS/MS techniques. Several other issues being addressed include recovery, carryover, ability of emitters to generate reproducible results after numerous sample injections and automation of the sample preparation. These issues in the context of CSF, plasma and brain sample preparation as well as nano-LC/MS method development will be the focus of this presentation. |
|
General Posters
7:00 PM-9:00 PM, Sunday, August 19, 2007 BCEC -- Exhibit Hall - B2, Poster
Sci-Mix
Division of Analytical Chemistry |