Abstract [Background] Although advanced imaging technology enables us to detect small breast lesions, it remains a challenge whether it is a benign or malignant tumor with imaging findings alone. A definitive diagnosis is required with cytological or pathological diagnosis under ultrasonography guidance. Excessive examinations and malpractice are also concerned with increased examinations. The purpose of the study was to identify the optimal diagnostic approach for small breast lesions.[Patients and Methods] We reviewed a total of 1532 cases of ultrasonography-guided vacuum-assisted core needle biopsy (VAB) performed at a single institution between June 1996 and December 2013. There were 519 small breast lesions (274 non-palpable lesions and 245 non-mass lesions). Ultrasonographic examinations were performed on a LOGIC 500 (GE Healthcare, Waukesha, WI, USA) using an 11 MHz linear transducer before November 2011, and on Aplio MX (TOSHIBA, Minato, Tokyo, Japan) using an 8 MHz linear transducer since then. We performed VAB under ultrasonography guidance using 11-gauge probes (Mammotome Biopsy system; Biopsys Medical Inc., Irvine, CA, USA) for diagnosis and 8-gauge probes for excision of the lesion.[Results] The mean age of the 519 patients was 52.7 years. Ultrasonography-guided fine needle aspiration cytology (FNAC) was performed before VAB in 269 cases (51.8%). The FNAC revealed 76 malignant, 23 suspected malignant, 92 indeterminate, 47 benign or normal, and 31 inadequate cases. Of these FNAC results, the final pathological diagnosis was benign in 2 of 76 malignant and 6 of 21 suspected malignant cases, and malignant in 4 of 47 benign cases. Accordingly, the true positive, true negative, false positive, and false negative rates for the 146 cases (excluding the indeterminate and inadequate cases) by FNAC were 95.8%, 84.3%, 15.7%, and 4.2%, respectively. The pathological results of the VAB specimens were malignant in 229 and benign in 290 cases. Of the 290 cases classified as benign by VAB, we performed post-VAB excisional biopsy in 20 cases. The reasons for this second pathological examination were malignant or suspicious findings for malignancy by FNAC (8 cases), inconsistent imaging and FNAC findings (5 cases), and others (7 cases). Excisional biopsy revealed malignancy in 3 of these 20 cases. Furthermore, we re-performed VAB in 9 out of the original 290 cases, and these were all diagnosed benign as with the first VAB. Excision of the lesion was the primary reason for the second VAB (6 cases), followed by inconsistencies with the imaging findings (2 cases), and suspected malignancy by FNAC (1 case). The true positive, true negative, false positive and false negative rates of VAB were 98.7%, 100%, 0%, and 1.3%, respectively. The mean follow-up duration was 43 months. [Conclusion] To prevent the excessive examinations, FNAC should be excluded from the initial diagnostic approach for small breast lesions. VAB is a highly reliable technique as the initial diagnosis for small breast lesions with high true positive and true negative rates and a very low false positive rate. The optimal strategy for diagnostic procedures should be adopted in consideration of reducing stress and anxiety in patients and costs. Citation Format: Satoko Nakano, Masahiko Otsuka, Akemi Mibu, Toshinori Oinuma. What is the optimal diagnostic approach for small breast lesions? Fine needle aspiration cytology vs. Vacuum-assisted core needle biopsy [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P5-03-02.