Various brittle fractures have been found to occur at grain boundaries in polycrystalline materials. In thin film interconnections used for semiconductor devices, open failures caused by electro- and strain-induced migrations have been found to be dominated by porous random grain boundaries that consist of a lot of defects. Therefore, it is very important to explicate the dominant factors of the strength of a grain boundary in polycrystalline materials for assuring the safe and reliable operation of various products. In this study, both electron back-scatter diffraction (EBSD) analysis and a micro tensile test in a scanning electron microscope was applied to copper thin film which is used for interconnection of semiconductor devices in order to clarify the relationship between the strength and the crystallinity of a grain and a grain boundary quantitatively. Image quality (IQ) value obtained from the EBSD analysis, which indicates the average sharpness of the diffraction pattern (Kikuchi pattern) was applied to the crystallinity analysis. This IQ value indicates the total density of defects such as vacancies, dislocations, impurities, and local strain, in other words, the order of atom arrangement in the observed area in nano-scale. In the micro tensile test system, stress-strain curves of a single crystal specimen and a bicrystal specimen was measured quantitatively. Both transgranular and intergranular fracture modes were observed in the tested specimens with different IQ values. Based to the results of these experiments, it was found that there is the critical IQ value at which the fracture mode of the bicrystal specimen changes from brittle intergranular fracture at a grain boundary to ductile transgranular fracture in a grain. The strength of a grain boundary increases monotonically with IQ value because of the increase in the total number of rigid atomic bonding. On the other hand, the strength of a grain decreases monotonically with the increase of IQ value because the increase in the order of atom arrangement accelerates the movement of dislocations. Finally, it was clarified that the strength of a grain boundary and a grain changes drastically as a strong function of their crystallinity.