Aims To investigate the use of a laser-based method of detection as a potential diagnostic test for the rapid identification of infectious agents in human blood. Methods and Results In this study, the successful differentiation of blood spiked with viruses, bacteria or protozoan parasites to clinically relevant levels is demonstrated using six blood types (O+, O-, AB+, A+, A-, B+) using blood from different individuals with blood samples prepared in two different laboratories. Experiments were performed using various compositions of filters, experimental set-ups and experimental parameters for spectral capture. Conclusions The potential for developing a laser-based diagnostic instrument to detect the presence of parasites, bacteria and viruses in human blood capable of providing analysis results within minutes was demonstrated. Significance and Impact of the Study There is an ongoing need for clinical diagnostics to adapt to newly emerging agents and to screen simultaneously for multiple infectious agents. A laser-based approach can achieve sensitive, multiplex detection with minimal sample preparation and provide rapid results (within minutes). These properties along with the flexibility to add new agent detection by simply adjusting the detection programming make it a promising tool for clinical diagnosis.