The review is devoted to measurement methods of bond rupture forces in complex biological molecules, namely, the unwinding forces of a DNA double helix. Mechanical methods not affecting electromagnetically a system under study, which is especially significant for biological systems, are considered. We describe two main methods: atomic force microscopy and rupture event scanning. The latter is a new method also based on the mechanical action but it has a much simpler instrumental implementation. The capabilities of both methods are compared and they are shown to be promising to investigate chemical bond rupture forces in biological systems. The application of these methods to study the strength of chemical bonds is associated with overcoming numerous technical difficulties in both performance of measurements themselves and chemical modification of conjugated surfaces. We demonstrate the applicability of these methods not only for fundamental studies of the strength of chemical bonds determining the stability and the related possibility of functioning of three-dimensional biomolecular complexes, but also for the design of biosensors based on the mechanical effect (quartz crystal microbalance, QCM), e.g., with an opportunity of rapid analysis of DNA.