Despite the introduction of routine vaccination against pertussis for more than a half century, leading to a drastic decline in the number of reported cases, pertussis continues to be an important respiratory disease afflicting unvaccinated infants and previously vaccinated children as well as adults in whom immunity has waned. The diagnosis of pertussis is challenging and accurate laboratory identification of Bordetella infections remains problematic. Common laboratory diagnostic methods used for pertussis diagnosis include culture, direct-fluorescent-antibody testing (DFA), serology and polymerase chain reaction (PCR). Culture of Bordetella pertussis is highly specific but fastidious and has limited sensitivity. DFA provides a much more rapid result, but has the disadvantage of poor sensitivity and specificity. Serology is not useful in infants. In older persons, it is hampered by the limitations of paired sera and it provides mainly a retrospective diagnosis. Such limitations of conventional diagnosis testing have led to the development of PCR assays. Notwithstanding its lack of standardization, PCR has been found to be more sensitive and more specific than other methods. In this report, we aimed to review current knowledge about the available diagnostic methods and tests that accurately diagnose pertussis.