Granulocyte colony-stimulating factor (G-CSF) is one of the most prominent endogenous proteins in broad clinical use. While its biological and clinical effects are relatively well studied, little is known about its endogenous formation in health and disease. However, such knowledge is crucial to decide in which situations G-CSF should be applied efficiently in the clinic, ie. when endogenous production does not suffice. The dramatic changes induced by G-CSF in the differential blood cell count are directly immunomodulatory, strengthening the innate defence by multiplying neutrophilic granulocytes. A multitude of further immunomodulatory effects contribute to the regulation of the concerted host defence. In this review, following a short introduction into the biology of G-CSF, the available data on endogenous formation in a number of animal models and human diseases is compiled. The cellular sources and inducers of G-CSF formation are reviewed and the regulation of G-CSF expression on both the transcriptional and translational level are discussed. The emerging understanding of the role and regulation of endogenous G-CSF formation opens up possibilities to define therapeutic windows as well as targets for diagnostics or drug development. Lastly, the modulation of G-CSF formation by various pharmacological agents alerts to putative side effects of these drug treatments.