<p>Background: The purpose of the study was to determine cut-off values for ovarian volume (OV) and follicle number per ovary (FNPO) in Indian women with polycystic ovary syndrome (PCOS).<br /> Methods: Eighty six PCOS women (Rotterdam criteria) and forty five matched ovulatory and normo-androgenic women were recruited. A detailed 2D and 3D trans-vaginal scan was carried out in early follicular phase (D2-D5) in all patients. Ovarian volume, follicle number per ovary, stromal volume, vascularization index (VI), vascularization flow index (VFI) and flow index (FI) were measured in PCOS and controls. Mann-Whitney test and logistic analysis using PROC LOGISTIC function of SAS^&reg; (Version 9.3) were used to calculate the best cut-offs for the diagnosis of PCOS.<br /> Results: Mean ovarian volume was 13.7&plusmn;5.89 and 5.06&plusmn;2.44 (p&lt;0.0001), FNPO was 19.18&plusmn;6.89 and 7.13&plusmn;3.51 (p&lt;0.0001) in PCOS and controls, respectively. The cut-offs for the diagnosis of PCOS were 2D OV=6.15 <em>cm</em>&sup3;, 2D FNPO=12. By 3D scan, OV=7 <em>cm</em>&sup3;, FNPO=10, stromal volume=6 <em>cm</em>&sup3;, VI=4.546, VFI=2.925 and FI=19.266. Youden&rsquo;s Index (To select optimal predicted probability cut-off) was the highest for 2D FNPO (0.88786). 2D FNPO showed the highest specificity and sensitivity (AUC), 0.95238 and 0.93548, for the diagnostic accuracy of PCOS.<br /> Conclusion: 2D and 3D trans-vaginal scans are equally accurate for assessment of ovarian morphology. FNPO has better diagnostic accuracy for PCOS compared to ovarian volume. Cut-off for FNPO and OV in Indian PCOS women is 12 and 6.15 <em>cm</em>&sup3; by 2D, 10 and 7 <em>cm</em>&sup3; by 3D trans-vaginal scan.</p>