Testicular microlithiasis (TM) is sometimes observed during scrotal ultrasound examinations in men. It has been suggested that TM is more prevalent in testes of men at risk for testicular carcinoma in situ (CIS), the precursor cells of all testicular germ cell tumours (TGCT). We have performed a retrospective analysis of ultrasound images and additional clinical data of a selected cohort of men and have determined the risk factor of TM and other ultrasound abnormalities for testicular CIS. Between 2002 and 2007, 176 testicular biopsies were performed in men with abnormalities found on the scrotal ultrasound. TM was found in 76/176 men (43.2%) and CIS was diagnosed in 20 of these men (26.3%). Here, we focused on the group of 76 men with TM to determine additional risk factors, besides TM, for CIS. In both groups, those with and without CIS, reproductive hormones, scrotal ultrasound images and patient history were compared. Predictive ultrasound findings for CIS were TM (sensitivity 100%, 95% CI: 0.8-1.0; specificity 64.1%, 95% CI: 0.6-0.7; PPV 26.3%, 95% CI: 0.2-0.4) and within this group an inhomogeneous testicular parenchyma (OR 16.1, 95% CI 2.4-106.8; sensitivity 75.0%, 95% CI: 0.5-0.9; specificity 79.0%, 95% CI: 0.7-0.9; PPV 50.0%, 95% CI: 0.3-0.7). Other significantly ultrasound characteristics for CIS in this population with TM were clusters of TM (p = 0.02) and intra-testicular lesions (p = 0.01). Men with CIS were found to have significantly lower values of inhibin-B (p = 0.02). Clusters of TM, intra-testicular lesions and lower values of inhibin-B were not significantly different in logistic regression analysis. TM on scrotal ultrasound of men with risk factors for TGCT and men with clinical signs of testicular maldevelopment has a high predictive value for CIS. However, the predictive value of an inhomogeneous testicular parenchyma, besides TM, for CIS is much higher.