In protic solution exchangeable protons such as those in hydroxyl or amine group exchange protons with the solvent. If DO is solvent, deuterons will be incorporated at these positions. The exchange reaction can be followed using a variety of methods (see Detection). Since this exchange is an equilibrium reaction, the molar amount of deuterium should be high compared to the exchangeable protons of the substrate. For instance, deuterium is added to a protein in HO by diluting the HO solution with DO (e.g. tenfold). Usually exchange is performed at physiological pH (7.0–8.0) where proteins are in their most native ensemble of conformational states.
The H/D exchange reaction can also be catalysed, by acid, base or metal catalysts such as platinum. For the backbone amide hydrogen atoms of proteins, the minimum exchange rate occurs at approximately pH 2.6, on average.  By performing the exchange at neutral pH and then rapidly changing the pH, the exchange rates of the backbone amide hydrogens can be dramatically slowed, or quenched.  The pH at which the reaction is quenched depends on the analysis method.  For detection by NMR, the pH may be moved to around 4.0–4.5.  For detection by mass spectrometry, the pH is dropped to the minimum of the exchange curve, pH 2.6. In the most basic experiment, the reaction is allowed to take place for a set time before it is quenched.
The deuteration pattern of a molecule that has undergone H/D exchange can be maintained in aprotic environments. However, some methods of deuteration analysis for molecules such as proteins, are performed in aqueous solution, which means that exchange will continue at a slow rate even after the reaction is quenched. Undesired deuterium-hydrogen exchange is referred to as back-exchange and various methods have been devised to correct for this.