This chapter describes the potential and limitations of attenuated total reflection infrared (ATR-IR) spectroscopy, modulation excitation spectroscopy (MES), and vibrational circular dichroism (VCD) for the investigation of bio-interfaces. The three techniques are completely independent of each other although ATR-IR and MES can be combined. ATR-IR is based on the total internal reflection of a light beam at an interface formed between two media. The resulting field at the interface is used for spectroscopy. The evanescent nature of this field allows one to probe the layer close to the interface formed between the internal reflection element (IRE) and the sample. Therefore, the field probes only a small part of the bulk fluid phase, such that molecules at the interface can be studied even in the presence of strongly absorbing solvents such as water. This makes the technique particularly interesting for applications in biology, where water is the solvent. The electric field can furthermore be polarized, which can be used for orientation measurements. MES is based on the stimulation of the sample under investigation by a periodic change of an external parameter such as temperature, pressure, pH, or concentration. The periodic system response to this stimulation (excitation) can be followed using a spectroscopic technique like ATR-IR by recording time-resolved spectra. The latter are then transformed into phase-resolved spectra using a phase-sensitive detection (PSD) scheme. MES leads to a drastic suppression of noise, which is very welcome when studying molecules at interfaces by spectroscopic techniques. Furthermore, MES helps to disentangle complex overlapping spectra arising from several species.