Blood is the most common and relevant bodily fluid that can be found in crime scenes. It is critical to correctly identify it, and to be able to differentiate it from other substances that may also appear at the crime scene. In this work, several stains of blood, chocolate, ketchup, and tomato sauce on five different substrates (plywood, metal, gauze, denim, and glass) were analysed by ATR FTIR spectroscopy assisted with orthogonal partial least square-discriminant analysis (OPLS-DA) models. It was possible to differentiate blood from the environmental interfering substances independently of the substrate they were on, and to differentiate bloodstains according to the substrate they were deposited on. These results represent a proof-of-concept that open new horizons to differentiate bloodstains from other interfering substances on common substrates present in crime scenes.