Nowadays the use of laser technology as a highly precise tool for the cleaning of paintings is gaining ground. The development of a non-invasive analytical protocol aimed at thorough assessment of the treated surfaces and real time monitoring of the laser cleaning action is thus becoming imperative. This ensures that no side effects (e.g. discoloration, darkening, blackening) will occur on the painting surfaces due to laser ablation. In the present study the potential of the combined use of optical coherence tomography (OCT) and reflection FT-IR spectroscopy for in situ non-invasive assessment of laser cleaning procedures has been investigated on a historical easel painting donated to science. Specifically, OCT and FT-IR analyses were carried out before and after each cleaning test in order to carefully assess the condition of the painting surfaces upon their irradiation with a KrF excimer laser and evaluate the removal of weathered and/or non-original materials (i.e. waxes, aged natural varnishes, oxalates, deposition/migration compounds). This specific laser radiation, operating in the ultraviolet region (λ = 248 nm), is highly absorbed by coating materials (i.e. oxidized natural varnishes) and thus high selectivity and control of the cleaning process can be pursued. A systematic integration of the information on the chemical changes of the molecular composition of the painting surface provided by reflection FT-IR spectroscopy with the visualization of the stratigraphical changes by OCT allowed the evaluation of the sensitivity of the painting materials to different operative parameters of the laser treatment, such as laser fluence and number of pulses. In parallel, laser induced fluorescence (LIF) spectroscopic measurements, recorded with the same laser beam at significantly attenuated energy density values, were exploited to provide information on the fluorescence properties of the irradiated varnish surfaces. The combination of the results obtained by OCT and FT-IR complemented with those from LIF measurements are discussed with the aim of developing an efficient methodology for assessing in situ the irradiated painting surfaces after the laser cleaning treatments and, thus, setting the basis for a future monitoring scheme that would ensure optimum cleaning interventions.