We prospectively determined the variability in radiologists' interpretation of screening mammograms and assessed the influence of type and number of readers on screening outcome. Twenty-one screening mammography radiographers and eight screening radiologists participated. A total of 106,093 screening mammograms were double-read by two radiographers and, in turn, by two radiologists. Initially, radiologists were blinded to the referral opinion of the radiographers. A woman was referred if she was considered positive at radiologist double-reading with consensus interpretation or referred after radiologist review of positive cases at radiographer double-reading. During 2-year follow-up, clinical data, breast imaging reports, biopsy results and breast surgery reports were collected of all women with a positive screening result from any reader. Single radiologist reading (I) resulted in a mean cancer detection rate of 4.64 per 1000 screens (95% confidence intervals (CI)=4.23-5.05) with individual variations from 3.44 (95% CI=2.30-4.58) to 5.04 (95% CI=3.81-6.27), and a sensitivity of 63.9% (95% CI=60.5-67.3), ranging from 51.5% (95% CI=39.6-63.3) to 75.0% (95% CI=65.3-84.7). Sensitivity at non-blinded, radiologist double-reading (II), radiologist double-reading followed by radiologist review of positive cases at radiographer double-reading (III), triple reading by one radiologist and two radiographers with referral of all positive readings (IV) and quadruple reading by two radiologists and two radiographers with referral of all positive readings (V) were as follows: 68.6% (95% CI=65.3-71.9) (II); 73.2% (95% CI=70.1-76.4) (III); 75.2% (95% CI=72.1-78.2) (IV), and 76.9% (95% CI=73.9-79.9) (V). We conclude that screener performance significantly varied at single-reading. Double-reading increased sensitivity by a relative 7.3%. When there is a shortage of screening radiologists, triple reading by one radiologist and two radiographers may replace radiologist double-reading.