Immunophenotyping provides valuable prognostic and diagnostic information, but is technically complex and expensive. The assessment of autofluorescence is label-free and provides complex information on cell identity. However, research on its application to immunophenotyping has been heterogenous. This systematic review was carried out to identify and synthesise all available evidence on the use of autofluorescence for immunophenotyping. Eighty three full texts were included. There was a focus on neutrophils (20 papers) and macrophages (22 papers) with alveolar macrophages (13 papers) forming a subcategory. Seven studies investigated monocytes, three focused on microglia, two on dendritic cells, five on mast cells, nine on granulocytes, thirteen on eosinophils, one on erythrophagocytic cells, and one on natural killer cells. Eleven studies investigated uncategorised immune cell populations. Translation of findings into clinical immunophenotyping requires the application of reproducible methods, along with clear reporting of excitation and emission parameters, and a greater focus on clinical and primary samples.