Among copepods, which are the most abundant animals on Earth, the genus Oithona is described as one of the most numerous and plays a major role in the marine food chain and biogeochemical cycles, particularly through the excretion of chitin-coated fecal pellets. Despite the morphology of several Oithona species is well known, knowledge of its internal anatomy and chitin distribution is still limited. To answer this problem, Oithona nana and O. similis individuals were stained by Wheat Germ Agglutinin-Fluorescein IsoThioCyanate (WGA-FITC) and DiAmidino-2-PhenylIndole (DAPI) for fluorescence microscopy observations. The image analyses allowed a new description of the organization and chitin content of the digestive and reproductive systems of Oithona male and female. Chitin microfibrils were found all along the digestive system from the stomach to the hindgut with a higher concentration at the peritrophic membrane of the anterior midgut. Several midgut shrinkages were observed and proposed to be involved in faecal pellet shaping and motion. Amorphous chitin structures were also found to be a major component of the ducts and seminal vesicles and receptacles. The rapid staining protocol we proposed allowed a new insight into the Oithona internal anatomy and highlighted the role of chitin in the digestion and reproduction. This method could be applied to a wide range of copepods in order to perform comparative anatomy analyses.