General sample preparation for 1.7mm tubes

The following guidelines will help with preparing 1.7mm tubes.

  • Small molecules need to be dissolved in a suitable high purity deuterated solvent: CDCL3, MeOD, D2O (99.96%).
  • Typically, only 30μL of sample is needed. Samples are loaded using a Hamilton syringe, capped with a Teflon ball to prevent evaporation and spun to remove air bubbles.
  • Proteins and peptides should be dissolved in a suitable buffer (10-50 mM) with 10% D2O. Ideally, we would aim for an acidic buffer (pH 4-7), but dissolve the sample in a buffer that provides maximum stability and solubility for your sample.
  • Salt may be need for protein/peptide solubility but be aware salty samples (>100mM NaCl) may reduce signal to noise on the cold NMR probe.
  • Typically, a few milligrams is sufficient to run the standard suite of experiments for a small molecule. Less sensitive experiments will require milligram quantities.
  • Structure determination by NMR typically requires a protein concentration of 0.5 mM or greater, stable for several days at the desired temperature. NMR studies for ligand binding or protein-protein interaction studies require concentrations of at least 0.1 mM.
  • Solid particles, especially those in cloudy solution drastically reduce spectral quality. Remove by centrifugation at high speed for at least 5 minutes.
  • Tubes should be check for damage/cracks and wiped with ethanol to remove dirt and grease before loading into the rack.
Left to right: 600μl in a 5mm tube. 200μl in a 3mm tube, 3mm shigemi tube with 70μl and 30μl in a 1.7mm tube. 1 pence piece to scale
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