Monolith NT.115

To ensure optimal results, please follow these rules when preparing the samples:

• Never prepare less than 20 µl of sample.

  • Increased probability of errors due to evaporation.

  • Sample material may stick to the plastic micro reaction tubes.

  • Higher pipetting errors.

• Never prepare small volumes (e.g. 20 µl) in large micro reaction tubes.

  • The high surface-to-volume ratio leads to surface adsorption.

  • Always use the smallest micro reaction tubes possible, like PCR tubes.

• Always spin down the protein/sample stocks (5 min at 13,000 rpm, 4°C).

  • This will remove aggregates (one of the main sources for noise).

  • In addition, use detergents such as Tween-20 or Triton X-100 (0.01% - 0.1%).

• Keep the samples in the dark once the fluorescently labeled molecule is added.

• Incubate the components of the binding reaction for a sufficient time to form interactions.

  • Incubation time and temperature can differ between different molecules and should be determined experimentally.

  • Usually, a 5-minute incubation at RT is sufficient.

• Avoid formation of bubbles during sample preparation.

  • It is recommended to mix the samples by pipetting rather than vortex these small volumes (vortex may lead to protein or sample denaturation).

• Use proper controls for the experiment.

  • A non-binding molecule with the molecule of interest.

  • A binding-deficient mutant of the molecule of interest.