or by pH changes (the i-motif requires a slightly acidic pH to form), and thus this molecular system is ideally suited to conceive control experiments. We studied here a 34mer C-rich sequence related to the promoter region of the n-myc oncogene (Figure , Table ). The wild type sequence shows an i-motif and an unusually long loop containing two complementary TGCA sequences which promote the formation of a stable hairpin. This sequence will be called IH (i-motif+hairpin). In addition, we studied 34mer control sequences, which can form only the i-motif (IC), only the hairpin (CH) and the last control, which can form none of the ordered structures (CC). Our goal was to establish whether CIU can be used to assess the preserved intramolecular folding of nucleic acids in electrospray conditions compatible with native-MS, with i-motifs and controls as model systems. To overcome the limitation of compaction at low charge states, we used supercharging agents in low amounts to promote native supercharging. We studied the resulting charge state distributions (CSD) and collision cross section distribution (CCSD) at low internal energy, and the CIU profiles of all charge states. We varied both the sequence (Figure , Table ) and the pH (at pH = 5.6 the i-motif is formed and at pH = 7.1 it is disrupted). A problem to be surmounted is that current software to describe unfolding processes in the gas phase, such as CUIsuite2 , allowed to distinguish automatically only two conformations in our molecular systems. To lift this limitation, we applied the method multivariate curve resolution-alternating least-squares to extract two or more physically meaningful pure components from the CIU data, according to the complexity of the unfolding procedure. Although we found evidence of a memory of the solution hydrogen bonds in the CSD, CCSD or CIU, we also uncovered several caveats in the conception of control experiments, and pitfalls in data interpretation.