The aims of this prospective study were to investigate the accuracy of B-mode transvaginal ultrasonography alone, using the typical finding of the presence of an elongated shaped mass with incomplete septa, in the screening of hydrosalpinx in women undergoing surgery for gynaecological diseases, and to determine the predictive value of this method combined with colour Doppler energy (CDE) imaging evaluation and CA125 concentrations in differentiating hydrosalpinx from other adnexal masses. In the first part of the study, 378 consecutive pre-menopausal non-pregnant women were submitted to transvaginal ultrasonography alone before surgery. In the second part of the study, 256 adnexal masses underwent transvaginal ultrasonography combined with CDE imaging evaluation associated with spectral Doppler analysis and plasma concentrations of CA125. Sensitivity and specificity for the ultrasonographic screening were 84.6 and 99.7% respectively, calculated for each adnexum (n = 756) and 93.3 and 99.6% respectively, calculated for each mass, for differentiating hydrosalpinx from other adnexal masses. The CDE imaging and the evaluation of CA125 plasma concentrations do not seem to increase the accuracy of B-mode transvaginal ultrasonography. Inter- and intra-observer agreement, expressed in terms of k-values, was high (0.87 and 0.93 respectively). In conclusion, transvaginal ultrasonography alone is a useful method of detection of hydrosalpinx.