Aptamers are single-stranded nucleic acids capable to bind selectively to a ligand or to a family of molecules. Aptamers are the sensing part of riboswitches, which are regulatory segments of messenger RNA involved in gene expression. Aptamers are also promising artificial probes, sensors and stimuli-responsive elements. In the development of aptamer-based technology, it is crucial to understand how binding is occurring, to quantify affinities, and ligand-induced conformational changes. The objective of this thesis is to explore the applicability of native IM-MS to DNA and RNA aptamers to quantify binding and to detect conformational change upon binding.In the first part, we evaluated the quantitative determination of equilibrium dissociation constants (KD) by mass spectrometry (MS), and the necessity of including a correction for relative response factors of free and bound aptamers. We compared isothermal titration calorimetry and MS titrations to validate the quantifications. Two RNA aptamers were taken as models: the malachite green aptamer, extensively studied by ITC, and the riboflavin mononucleotide aptamer, a case of Mg2+-dependent ligand binding. We observed that typical volatile electrolytes ammonium acetate and trimethyl ammonium acetate are suitable to study RNA aptamer binding, and that comparable KD values are obtained from ITC and native MS. The neomycin and tobramycin RNA aptamers were chosen to test the limit of detection of native MS. We found that native MS is appropriate to determine KD values in the range from 50 nM to 30 µM. The relative response factor correction was relatively modest in all cases, suggesting that the ligand binding is not associated to a significant conformational difference upon ionization. For these aptamers, we conclude that assuming equal response factors is acceptable.In the second part, we evaluated whether the aptamers’ “adaptive binding” mechanism can be revealed by ion mobility spectrometry (IMS). To this aim, in addition to the systems listed above we studied the tetracycline RNA aptamer and a series of cocaine-binding DNA aptamers, for which the conformational change upon binding is reported in literature. For all aptamers except the tetracycline aptamer, we did not observe a significant difference in the shape of the gas-phase structure upon ligand or Mg2+ binding. However, a significant change was observed in tetracycline RNA aptamer’s ion mobilities, at biologically relevant concentration of Mg2+ (100 µM), and we found that Mg2+ is essential for ligand binding, in agreement with previous solution studies. For the cocaine-binding DNA aptamer series, although we observed similar compactness for the free and bound aptamers in soft pre-IMS conditions, a conformational extension occurs at high pre-IMS activation, best revealed by charge state 7-, suggesting gas-phase rearrangements. To better investigate whether the energetics of these rearrangements depend on pre-folding or on ligand binding, we modified the sequences with dA overhangs, to compare systems with similar numbers of degrees of freedom without altering the core structure. We also propose new ways of presenting the data, adapted to the cases where ligand dissociation, declustering and unfolding occur at similar voltages. The gradual increase of the pre-IMS collisional activation revealed that the unfolding energetics is correlated with the base pairs content, suggesting that base pairs are conserved in the gas-phase structures. We also found that ligand is lost at lower energies than unfolding.In summary, gas-phase compaction occur for both the free aptamers and bound aptamers, and memories of the solution-phase structures can only be revealed in some particular cases. However, the compaction towards similar shapes might constitute an advantage for the quantification, because molecular systems of similar shapes have similar electrospray responses. Consequently, native MS provides reliable estimations of KD values.