Research and development (R&D), especially in industry and technology, is a crucial component of innovation, productivity and effectiveness, as well as the generation of new competitive advantages that are reflected in the whole economy of a country. In this context and with the aim of improving the measurement of R&D, the Organization for Economic Cooperation and Development updated the Frascati Manual in 2015 to provide the main, globally applicable guidelines for national statistical offices and policy-makers to analyze trends and allow for international comparisons of science, technology and innovation. In developing countries, it is particularly important to determine different strategies to guarantee adequate R&D to inform the creation of policies and instruments to effectively promote knowledge and generate technological solutions to local problems. This chapter describes the development and application of the Frascati Manual (2015) in Colombia to measure investments in R&D and other scientific, technological and innovation activities in Colombia from the perspective of the requirements of the manual for government, higher education, business enterprises, the health sector and private non-profit institutions with an update for the entire 2000–2017 period. The results indicate that the new guidelines for quantifying investments in R&D guarantee greater reliability through the use of mixed methods involving administrative registers or surveys and control mechanisms with budget analysis. Moreover, it is important to analyze differences among sectors to adequately determine the specific factors related to scientific, technological and innovation activities with the aim of establishing the investments and expenditures in science, technology and innovation. During the period of study, Colombia maintained constant investment with a slight increase in the last year that was mainly due to found royalties. The resulting indicators of expenditure in R&D and other scient.