Background: Many factors affecting satellitism test are not clear, and it is difficult to avoid misidentified even if the medium is properly selected. We investigated the factors that cause false positives of Haemophilus influenzae and false negatives of Haemophilus parainfluenzae in the satellitism test supplemented V factor by Staphylococcus aureus.Methods: H. influenzae (four reference strains and 47 clinical isolates), H. parainfluenzae (two reference strains and 67 clinical isolates), four different media, and two strains of S. aureus activated on two different media were involved in this study.Results: The type of medium used to activate S. aureus was the most common factor causing false positives of H. influenzae, followed by the strain of S. aureus, and again the type of medium used for the experiment. The production of false negatives of H. parainfluenzae was only related to the medium used in the experiment.Conclusions: Tryptic soy agar and S. aureus (ATCC 25923) activated with nutrient agar should be used in the satellitism test for Haemophilus spp. to improve the accuracy of the test.