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In this work, ultraviolet-visible reflection spectroscopy is proposed as a technique that, in combination with classical surface pressure–area isotherms, allows to study in situ the adsorption of DNA to octadecylamine monolayers. The presence of the polynucleotide molecules at the interface, typically demonstrated by an expansion and a change in the profile of the ODA isotherms, has been confirmed here by a reflection peak at 260 nm. Increasing DNA concentrations in the subphase from 2 to 8 μM is accompanied by an increment in the expansion of the isotherm showing in all cases an abrupt phase transitions at high surface pressures that is also observed in lateral compressibility representations. This phase transition has been attributed to a squeezing out of DNA phenomenon as demonstrated by the normalization of the corresponding reflection spectra. In addition, hysteresis and reversibility in the formation of the monolayer has been identified, making possible the realization of successive compression–decompression cycles without reaching the collapse of the monolayer. It can be inferred that the DNA molecules expelled out of the monolayer at high surface pressures can enter again during the decompression process. Reflection spectroscopy has been also found to be a valuable tool to investigate the dependence of the adsorption process with both time and DNA concentration in the subphase.

Unveiling the interaction of DNA–octadecylamine at the air–water interface by ultraviolet-visible reflection spectroscopy