Serum Preparation for 1D NMR: Difference between revisions
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8. At this point, the supernatant samples are collected and snap frozen in Liquid Nitrogen and freeze dried to remove methanol by speed vacuum centrifugation (SpeedVac R Plus, Savant) and water by lyophilization using FreeZone™ (Labconco, Kansas City, MO) for at least 24 hours and stored at -20°C until NMR analysis. | 8. At this point, the supernatant samples are collected and snap frozen in Liquid Nitrogen and freeze dried to remove methanol by speed vacuum centrifugation (SpeedVac R Plus, Savant) and water by lyophilization using FreeZone™ (Labconco, Kansas City, MO) for at least 24 hours and stored at -20°C until NMR analysis. | ||
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Revision as of 04:01, 11 September 2021
1. Serum sample are first aliquoted
2. Proteins are precipitated by the addition of twice the amount of methanol as sample
3. Antibacterial NaN3 is added 2.0% by volume to avoid any bacterial growth
4. The 1:2 mixture of serum: solvent is then vortexed for 10 seconds
5. Samples are incubated at 4°C for 10 minutes
6. Next, the samples are centrifuged at 14,000 rpm for 20 mins at 4°C to pellet the proteins
7. The supernatant is collected and centrifuged at 14,000 rpm for 5 min a 4°C to pellet any leftover protein
8. At this point, the supernatant samples are collected and snap frozen in Liquid Nitrogen and freeze dried to remove methanol by speed vacuum centrifugation (SpeedVac R Plus, Savant) and water by lyophilization using FreeZone™ (Labconco, Kansas City, MO) for at least 24 hours and stored at -20°C until NMR analysis.