• Suitable sample preparation methods, probe types, and imaging modes are recommended for AFM-IR tests. • Some imaging parameters have the significant effect on imaging quality. • The difference of nanoscale property of various phases on bitumen surface may not be entirely caused by chemical composition. Atomic force microscopy-based infrared spectroscopy (AFM-IR) combines the chemical analysis capability of IR with the nanoscale resolution of AFM. Considering the importance of test methods to imaging quality, it is necessary to explore the suitable AFM-IR test methods for bitumen materials including bitumen and bituminous mixtures. As for bitumen sample preparation, solution-cast methods, heat-cast methods, and hot-bitumen-pouring methods have proved to be feasible. Additionally, this study develops frozen-storage and low-temperature-cutting methods to prepare bituminous mixture samples for AFM-IR tests. Afterwards, samples of base bitumen aged by PAV for 25 h are subjected to explore the influence of different probe types, imaging modes, and test parameters. It is found that the PR-EX-TnIR-A probe can be used to capture chemical information in the tapping mode and the contact mode. Accordingly, only the tapping mode is suitable for obtaining rapid narrow-band chemical maps, while the contact mode can be used to collect broad-band point infrared spectra besides the tapping mode. On this basis, the variation of test parameters has shown that IR power and start power strongly influence the obtained chemical information. Finally, based on the proposed AFM-IR test methods for bitumen materials, we capture infrared spectra of each phase on the virgin base bitumen surface and then conduct the analysis of surface phase characteristics, combining with nanomechanical properties obtained by the AFM peak force quantitative nanomechanical mode (PF-QNM). The results indicate that chemical composition of various phases is similar, and the obvious difference of microstructure and nanomechanical properties is not completely related to chemical composition.