As state-of-the-art electrochemical energy conversion and storage (EECS) techniques, fuel cells and rechargeable batteries have achieved great success in the past decades. However, modern societies’ ever-growing demand in energy calls for EECS devices with high efficiency and enhanced performance, which mainly rely on the rational design of catalysts, electrode materials, and electrode/electrolyte interfaces in EESC, based on in-deep and comprehensive mechanistic understanding of the relevant electrochemical redox reactions. Such an understanding can be realized by monitoring the dynamic redox reaction processes under realistic operation conditions using in situ techniques, such as in situ Raman, Fourier transform infrared (FTIR), and X-ray diffraction (XRD) spectroscopy. These techniques can provide characteristic spectroscopic information of molecules and/or crystals, which are sensitive to structure/phase changes resulted from different electrochemical working conditions, hence allowing for intermediates identification and mechanisms understanding. This review described and summarized recent progress in the in situ studies of fuel cells and rechargeable batteries via Raman, FTIR, and XRD spectroscopy. The applications of these in situ techniques on typical electrocatalytic electrooxidation reaction and oxygen reduction reaction (ORR) in fuel cells, on representative high capacity and/or resource abundance cathodes and anodes, and on the solid electrolyte interface (SEI) in rechargeable batteries are discussed. We discuss how these techniques promote the development of novel EECS systems and highlight their critical importance in future EECS research.