Assessment of Fine-Needle Aspiration Sampling Technique in Thyroid Nodules Yolanda M. Musgrave, M.D., 1 Diane D. Davey, M.D., 1 * Julia A. Weeks, M.D., 1 Evelyn R. Banks, M.D., 1 Mary K. Rayens, Ph.D., 2 and Kenneth B. Ain, M.D. 3,4 BACKGROUND: A prospective two-yr study was undertaken to assess the heterogentity of thyroid nodules using the fine-needle aspiration (FNA) technique of systematic regional sampling. In addition, we determined the number of regions to be sampled to minimize non-diagnostic results, and to optimally characterize thyroid nodules. DESIGN: FNA was performed on 74 nodules  1.5 cm. in diameter in five distinct regions in sequence (center, then four quadrants starting at 12:00, clockwise). Slides from each region were coded, randomized, subjected to blind review, and catego- rized as non-diagnostic (ND), benign (B), indeterminate (ID), suspicious/neoplastic (S/N), or malignant (M). Final cytologic diagnosis (CD) was made from all slides of each nodule. RESULTS: The ND rate for center FNAs alone was 16%, but addition of the 12:00 region decreased it to 5.3%. With 3, 4, or 5 sequential sites the nondiagnostic rates were 4, 2.6, and 2.6%. The center region diagnosis was identical to the final CD in 71% of the cases. Addition of the 12:00 region increased the concordance to 88%. Three sequential regions equaled the CD in 93% of cases, and 4 regions equaled the CD in 99% of cases. All nodules characterized as M or S/N were resected as were 76% of the ID nodules. Of the 43 nodules characterized as B, 3 were resected, 24 involuted, 6 were unchanged, and 10 were lost to follow-up. All 3 M nodules proved malignant by histology, as did 7/10 S/N, 0/17 ID, and 0/43 B nodules; 3/10 S/N, 1/17 ID and 1/43 B were adenomas. Likelihood ratios for diagnosing neoplasia were ND:0, B:0.10, ID:0.21, S/N:, M:. CONCLUSIONS: Sampling of at least four distinct regions accurately assesses thyroid nodules while minimizing ND results. Regional sampling also addresses intranodular heterogeneity. Diagn. Cytopathol. 1998;18:76–80.  1998 Wiley-Liss, Inc. Key Words: thyroid; fine-needle aspiration; biopsy; sampling; likelihood ratios Although many techniques for performing fine-needle aspi- ration (FNA) of thyroid nodules have been described, 1–5 few prospective studies have validated these approaches. We undertook a 2-yr prospective study to ascertain cytologic heterogeneity of thyroid nodules by systematic geographic sampling. In addition, we determined the number of regions to be sampled to minimize non-diagnostic results and to adequately assess thyroid nodules. Diagnostic accuracy was assessed using likelihood ratio methodology. Materials and Methods Between September 1991 and September 1993, 75 FNAs were performed on 74 thyroid nodules (one nodule was aspirated twice in an interval of 17 mo). The nodules ranged in size from 1.5 to 8 cm. Cytologic material was procured by a single operator (K.B.A.) using a 27-gauge needle attached to a 10-ml syringe in a plastic aspiration holder. The nodules were aspirated in a systematic and sequential fashion starting at the central region and proceeding to four quad- rants (12, 3, 6, and 9 o’clock). The material from each pass was expressed onto slides that were labeled to identify the specific region as well as the patient. Half of the slides were immediately spray-fixed with cytology fixative for Papanico- laou staining, and the other half were air dried and stained by Diff-Quik method. Each pass yielded between 2 to 10 slides. If cyst fluid was obtained in the first pass, the liquid contents of the cyst were evacuated and the residual mass was aspirated as previously described. Slides from each of the 375 regions were separated, the labels were masked, and the slides were relabeled with numbers obtained from a random number generator. The sets of slides from each region were then randomized and 1 Department of Pathology and Laboratory Medicine, Markey Cancer Center, University of Kentucky, Lexington, Kentucky 2 Section of Statistics, Markey Cancer Center, University of Kentucky, Lexington, Kentucky 3 Medical Service, VeteransAffairs Medical Center, Lexington, Kentucky 4 Department of Internal Medicine, University of Kentucky Medical Center, Lexington, Kentucky Presented, in part, at the 84th Annual Meeting of the United States- Canadian Academy of Pathology, Washington, DC, 1995. Grant sponsor: NCI; Grant number: CA58935; Grant sponsor: VA Merit Review; Grant number: 93-006V; Grant sponsor: Ephraim McDowell Cancer Research Foundation, Lexington, KY; Grant sponsor: Lexington Clinic Foundation, Lexington, KY. *Correspondence to: Diane D. Davey, M.D., Department of Pathology and Laboratory Medicine, University of Kentucky Medical Center, 800 Rose Street, Room MS117, Lexington, KY 40536-0084. Received 26 June 1996; Accepted 7 May 1997 76 Diagnostic Cytopathology, Vol 18, No 1  1998 WILEY-LISS, INC.