Testicular microlithiasis (TM) is an unusual ultrasonographic manifestation in testicular parenchyma. Limited information is available about TM in Taiwanese men. We performed a retrospective analysis to investigate the characteristics of TM and its association with testicular cancer and infertility in Taiwan. Male patients who had received scrotal ultrasonography because of scrotal symptoms or infertility between January 2000 and December 2003 were recruited. The incidence of TM was 7.6%. Both testicular microlithiasis and testicular cancer occurred chiefly in the third decade. Patients with TM exhibit a higher chance of testicular cancer (6% vs. 0.9%). No local field effect between TM and testicular cancer was observed. Testicular microlithiasis severity is not positively correlated with sperm quality and sterility. Forty-eight patients (32%) were available at follow-up. No patient developed a testicular tumor or elevated tumor markers (AFP, beta-hCG) during follow-up. We suggest monthly self-examination, annual scrotal ultrasonography and tumor markers screening between the age of 20 and 30 years of patients with TM.