Biological functions essentially consist of a series of chemical reactions, including intermolecular interactions, and also involve the cooperation of a number of biological molecules performing these reactions. To understand this function at the molecular level, all steps of the reactions must be elucidated. However, since the biosystems including the surrounding environment are notably large, the reactions have to be elucidated from several different approaches. A variety of techniques have been developed to obtain structural information, and the knowledge of the three-dimensional structure of biomolecules has increased dramatically. Contrarily, the current information on reaction dynamics, which is essential for understanding reactions, is still not enough. Although frequently used techniques, such as spectroscopy, have revealed several important processes of reactions, there are various hidden dynamics that are not detected by these methods (silent dynamics). For example, although water molecules are essential for bioreactions, dynamics of the protein-water interaction are very difficult to trace and spectrally silent. Transient association/dissociations of proteins with partner proteins are difficult to observe. Another important property to understand the reaction of proteins is fluctuations, which are random movements that do not change the average structure and energy. The importance of fluctuations has been pointed out in order to explain enzymatic activity; however, it is extremely difficult to detect changes in fluctuation during a reaction. In this Account, unique time-resolved methods, time-resolved thermodynamics, and time-resolved diffusion methods, both of which are able to detect silent dynamics in solution at physiological temperature, are described.Thermodynamic properties are important for characterizing materials, in particular, macromolecules such as biomolecules. Therefore, the data available regarding these properties, for several stable proteins, is abundant. However, it is almost impossible to characterize short-lived intermediate species in irreversible reactions using traditional thermodynamic techniques. Similarly, although the translational diffusion coefficient is a useful property to determine the protein size and intermolecular interactions, there have been no reports revealing reaction dynamics. The transient grating (TG) method enables us to measure these quantities in a time-resolved manner for a variety of irreversible reactions. With this method, it is now possible to study biomolecule reactions from the viewpoint of thermodynamic properties and diffusion, and to elucidate reaction dynamics that cannot be detected by other spectroscopic methods.Here, the principles of the methodologies used, their characteristic advantages, and their applications to protein reactions are described. The TG measurements of octopus rhodopsin revealed a spectrally hidden intermediate and determined an energetic profile along the reaction coordinate. This emphasizes that the measurement in solution, not for trapped intermediates, is important to characterize the reaction intermediates. The application of these methods to a blue light sensor PixD revealed many spectrally silent dynamics as well as the importance of fluctuation for the reaction. As an example of the time-resolved heat capacity change and transient thermal expansion measurements, the reaction of PYP was briefly described. The reaction scheme of another blue light sensor protein, phototropins, and a spectrally silent DNA binding process of EL222 were fully elucidated by the time-resolved diffusion method.