Following a request from the European Commission, the EFSA Panel on Nutrition, Novel Foods and Food Allergens (NDA) was asked to deliver an opinion on 2'-fucosyllactose/difucosyllactose (2'-FL/DFL) mixture as a novel food (NF) pursuant to Regulation (EU) 2015/2283. The NF is a powdered mixture mainly composed of two oligosaccharides, 2'-FL and DFL, which are produced together by fermentation with a genetically modified strain of Escherichia coli K12. The information provided on the manufacturing process, composition and specifications of the NF does not raise safety concerns. The applicant intends to add the NF in a variety of foods, including infant and follow-on formula, foods for infants and young children, foods for special medical purposes and food supplements. The target population is the general population except for food supplements, for which the target population is individuals above 1 year of age. Since the intake of 2'-FL and DFL from the NF at the proposed use levels is unlikely to exceed the intake level of naturally occurring 2'-FL and DFL in breastfed infants per kilogram body weight, the Panel concludes that the NF, a mixture of 2'-FL and DFL, is safe under the proposed conditions of use for the proposed target population.