TA Instruments Nano ITC Series, Getting Started Manual

Page: 27 / 46

Nano ITC Getting Started Guide Page 27
Preparing and Degassing the Solutions
Large sample molecules are often stored in buffer which will affect the final pH of the prepared solution.
Dialysis is the process used to equalize the solution characteristics while retaining the large molecules.
This step improves the experimental results by minimizing the enthalpies of dilution and neutralization.
Dialyze the samples in the buffer, when possible, to minimize blank effects. All solutions used in the
experiment (rinse buffer, sample titrant, and sample titrand) must be degassed prior to use. When preparing
solutions for use with the Nano ITC, any solutions containing buffers and macromolecules should be dia-
lyzed before use, if possible. This is a standard process that is used to equalize the pH and concentration
between the sample titrant and the sample titrand.
Follow these steps to prepare the solutions:
1 Prepare a large amount of buffer as appropriate for the experiment (add the appropriate type and amount
of salts). This buffer will be used as material for formulation of the titrant and titrand solutions, and in
their dialysis.
2 Formulate solutions of any large-molecule sample compounds at this time using the buffer solution.
(Small-molecule sample compounds will be made up in a following step.)
3 Dialyze the solution(s) inside the remaining buffer. Place the sample in a dialysis bag and suspend it
inside the buffer solution. Gently stir the buffer for several hours to aid in the equalization of pH and
the concentrations of electrolytes. Temperature-sensitive samples may need to remain chilled during
this process.
4 Small molecule samples are prepared at this time using the dialyzed buffer. Do not dialyze this solution.
5 Retain 50–300 mL of the dialyzed buffer for use later in cell rinsing and for optional blank experiments.
Degassing Solutions
Typically, if a solution is heated, gas bubbles will form as the solubility of dissolved gases (e.g., O
2
and
N
2
) is decreased with increasing temperature. If gas bubble formation occurs in the ITC cells during the
run, the resulting data will be rather noisy since abrupt changes will result from the bubble-driven liquid
displacement effects.
All solvents must be degassed prior to being placed in the ITC to minimize the possibility of gas bubble
formation during the run. Pull a vacuum of 0.3–0.5 atm on the solutions for a period of 10–15 min to degas
a sample.
An accessory degassing system is available from TA Instruments.