Formamide/water mixtures were used as subphase for a Langmuir film of an amphiphilic iron(II) complex exhibiting a thermally-induced spin-crossover phenomenon around room temperature. The use of formamide enables the chemical stabilization of this complex (contrary to a pure water subphase) and leads to perfectly defined Langmuir-Blodgett (LB) films of this molecule. Variable temperature infrared spectrometry demonstrates that the low-spin to high-spin conversion occurs in this multilayer. Such a strategy based on a monolayer spread on organic solvent permits the formation of Langmuir and LB films from compounds chemically sensitive to water.