Chlamydia antibody testing (CAT) by micro-immunofluorescence (MIF) tests has been introduced into the fertility work-up as a screening test for tubal factor subfertility. In this study the role of C. pneumoniae antibodies, as a cause for false positive CAT results due to cross-reactivity with C, trachomatis antibodies in the MIF test, has been evaluated. In 240 subfertile women serological data were compared to laparoscopy findings, The prevalence of C, pneumoniae antibodies using enzyme-linked immunosorbent assay (ELISA) was 75% and did not differ between patients with and without tubal pathology, C, pneumoniae antibodies were found in 87% of women with a positive MIF test (greater than or equal to 32), and in 66% with a negative MIF test (P < 0.0005). Using ELISA instead of MIF for the detection of C. trachomatis antibodies, C, pneumoniae antibodies were found in 87% of C. trachomatis positive women, and in 69% of C, trachomatis negative women (P < 0.0005). Patients without tubal factor subfertility but a positive MIF test showed C, pneumoniae antibodies more frequently than patients without tubal factor subfertility and a negative MTF test. Therefore it was suggested that C, pneumoniae antibodies may be the cause of false positive CAT results. Remarkably, tubal pathology was more common in patients who had antibodies to both C. trachomatis and C, pneumoniae.