Chlamydiae are obligate intracellular bacteria. Chlamydia trachomatis is the most common sexually transmitted disease (STD). The C. trachomatis damaging disease sequelae such as sterility is based on intense and chronic inflammation elicited and maintained by reinfection or persistent infection. The high prevalence of C. trachomatis infection reflects the long and successful adaptation of these organisms to persist in their human host population. The large group of asymptomatically infected persons is not only at risk of serious long-term sequelae but also sustains transmission within communities. C. trachomatis acute infections have been diagnosed by cell culture, direct immunofluorescence, enzyme immunoassay, direct DNA hybridization, and more recently by nucleic acid amplification tests (NAATs). In chronic or persistent chlamydial infections, the level of Chlamydia is very low and bacteria are often not viable. Such infections would be characterized by continuing positive NAATs but only intermittent isolation of viable Chlamydia and positive assays for chlamydial protein antigen. The development of NAATs has been a major advance in the field of chlamydial diagnosis. The use of NAATs associated with serology test is the best diagnosis. The introduction of assays based on amplification of genetic material has subsequently increased the sensitivity of detecting chlamydial infections and offers the opportunity to use non-invasive sampling techniques to screen for infections in asymptomatic subjects. In this article, it was proposed the best diagnosis approaches for detection of acute and chronic infections.