Background: To evaluate knowledge about screening tests and tests without proven screening value in a Greek Breast Unit population undergoing benign vacuum-assisted breast biopsy (VABB). Materials and Methods: This study included 81 patients. Three knowledge-oriented items (recommended or not, screening frequency, age of onset) were assessed. Regarding screening tests two levels of knowledge were evaluated: i). crude knowledge (CK), i.e. knowledge that the test is recommended and ii). advanced knowledge (AK), i.e. correct response to all three knowledge-oriented items. Solely CK was evaluated for tests without proven screening value. Risk factors for lack of knowledge were assessed with multivariate logistic regression. A second questionnaire was administered 18 months after VABB to assess its impact on the performance of tests. Results: Concerning screening tests considerable lack of AK was noted (mammogram, 60.5%; Pap smear, 59.3%; fecal occult blood testing, 93.8%; sigmoidoscopy, 95.1%). Similarly lack of CK was documented regarding tests without proven screening value (breast self-examination, 92.6%; breast MRI, 60.5%; abdominal ultrasound, 71.6%; barium meal, 48.1%; urine analysis, 90.1%; chest X-Ray, 69.1%; electrocardiogram, 74.1%; cardiac ultrasound, 75.3%). Risk factors for lack of AK were: place of residence (mammogram), age (Pap smear), personal income (sigmoidoscopy); risk factors for lack of CK included number of offspring (breast MRI, chest X-Ray), BMI (abdominal ultrasound), marital status (urine analysis), current smoking status (electrocardiogram). VABB's only effect was improvement in mammogram rates. Conclusions: A considerable lack of knowledge concerning screening tests and misperceptions regarding those without proven value was documented.