From Fine-needle Aspiration Biopsy Research Articles

Content of RNA originating from thyroid in washouts from fine-needle and core-needle aspiration biopsy - preliminary study.

In the evaluation of molecular markers in washouts from fine-needle aspiration biopsy (FNAB) the extremely small amount of material can be a major problem. Some authors tried to use washouts from core-needle aspiration biopsy (CNABs) to gain more material from larger needles. However, according to some studies, CNAB samples are commonly contaminated with blood. The aim of our study was to evaluate the proportion of nucleic acids from thyroid cells in washouts from FNAB and CNAB by measuring the relative expression of cytokeratin 17 (KRT17) on the mRNA level. Relative expression of KRT17 and GADPH (reference gene) in washouts from FNAB and CNAB was measured using real-time PCR technique and compared to the results from surgical specimens. Surgical specimens form 22 nodules, FNAB samples from 20 lesions and CNAB samples from 24 lesions were analysed. The median difference in cycle threshold (Ct) between FNAB samples and surgical specimens was 3.3 (p = 0.047). In CNAB samples KRT17 was undetectable in most cases (median incalculable; proportion of samples with undetectable KRT17 significantly higher than in FNAB samples). Samples obtained with different biopsy techniques had different proportions of contents. The proportionally low content of epithelial cells in CNAB can result in underestimated expression of molecular markers of malignancy. Consequently, the risk of malignancy or unfavourable prognosis can also be underestimated. To conclude, results obtained from samples gained with one biopsy technique cannot be directly related to thresholds, and generally with experiences gained with other techniques, because it can lead to incorrect clinical interpretation of the results. (Endokrynol Pol 2016; 67 (6): 550-553).

Suspicious for follicular neoplasm or follicular neoplasm? The dilemma of a pathologist and a surgeon.

Cytological material obtained from Fine Needle Aspiration Biopsy (FNAB) does not permit us to distinguish between follicular carcinomas, adenomas, and hyperplastic nodules. The limitations of the method are: lack of possibility to assess the presence of tumour capsule, eventual capsular invasion, and angioinvasion. An unequivocal conclusion of whether what we have to deal with is a neoplastic or benign lesion is possible only after histopathological examination. The aim of the study was to confirm justification for using the term "Suspicious for Follicular Neoplasm" (SFN) in cytological diagnostics of thyroid carcinoma. Three hundred and fifty-two primary SFN FNAB diagnoses (diagnostic category IV [DC IV] - according to Bethesda System) obtained from 2010 to 2015 in the Institute of Oncology in Gliwice were analysed, and their correlation with histopathological diagnoses was verified. In the Institute of Oncology in Gliwice, 352 primary SFN diagnoses (diagnostic category IV [DC IV] - according to Bethesda System) were established. Surgical treatment was undertaken after first FNAB in six cases, giving confirmation of a neoplasm in five cases, one of which was a follicular carcinoma. Second FNAB performed in 90 patients confirmed DC IV diagnosis in 53 cases. Third FNAB concerned 26 patients, providing another 14 diagnoses of DC IV. 26 out of 352 patients were subjected to surgery, and then histopathological examination confirmed a neoplasm in 19 cases (which comprises 73%), five of which were carcinomas. High positive predictive value PPV = 73% of SFN diagnosis justifies undertaking surgical treatment in any case of this diagnosis.

Biopsy techniques for intraocular tumors.

Biopsy involves the surgical removal of a tissue specimen for histopathologic evaluation. Most intraocular tumors are reliably diagnosed based on the clinical evaluation or with noninvasive diagnostic techniques. However, accurately diagnosing a small percentage of tumors can be challenging. A tissue biopsy is thus needed to establish a definitive diagnosis and plan the requisite treatment. From fine-needle aspiration biopsy (FNAB) to surgical excision, all tissue collection techniques have been studied in the literature. Each technique has its indications and limitations. FNAB has been reported to provide for 88–95% reliable and safe ophthalmic tumor diagnosis and has gained popularity for prognostic purposes and providing eye conserving treatment surgeries. The technique and instrumentation for biopsy vary depending upon the tissue involved (retina, choroid, subretinal space, vitreous, and aqueous), suspected diagnosis, size, location, associated retinal detachment, and clarity of the media. The cytopathologist confers a very important role in diagnosis and their assistance plays a key role in managing and planning the treatment for malignancies.

Improved DNA Extraction Method for Molecular Diagnosis from Smaller numbers of Cells

Isolating total DNA from small samples using traditional methods is difficult and inefficient mainly due to loss of DNA during filtration and precipitation. With advances in molecular pathology, DNA extraction from micro-dissected cells has become essential in handling clinical samples. Genomic DNA extraction using small numbers of cells can be very important to successfully PCR amplify DNA from small biopsy specimens. We compared our experimental genomic DNA extraction method (A) with two other commercially available methods: using spin columns (B), and conventional resins (C), and determined the efficacy of DNA extraction from small numbers of cells smeared on a glass slide. Approximately 50, 100, 200, 500 and 1000 cells were isolated from fine needle aspiration biopsy (FNAB) slides aspirated from histologically proven papillary thyroid carcinoma masses. DNA was extracted using the three techniques. After measuring DNA quantity, PCR amplification was performed to detect the b-globin and BRAF V600E gene mutations. DNA extracted by method (A) showed better yield than the other methods in all cell groups. With our method, a suitable amount of genomic DNA to produce amplification was extracted from as few as 50 cells, while more than 100 to 200 cells were required when methods (B) or (C) were applied. Our genomic DNA extraction method provides high quality and improved yields for molecular analysis. It will be especially useful for paucicellular clinical samples which molecular pathologists often confront when handling fine needle aspiration cytology, exfoliative cytology and small biopsy specimens.

Thyroid Lesions Visualized on CT: Sonographic and Pathologic Correlation

There are no definitive guidelines for the management of incidental thyroid lesions on computed tomography (CT). The objectives of our study were to assess the association between CT and ultrasound (US) characteristics of thyroid lesions and identify CT predictors of benignity or malignancy. Two hundred fifty-nine patients who had undergone both CT of the chest or neck and thyroid US had at least one thyroid lesion visible on CT; both incidentally detected and palpable or symptomatic lesions were included. The CT and US examinations were retrospectively reviewed and lesions characterized. Pathologic results from fine-needle aspiration (FNA) and surgical excision were used to classify lesions as benign or malignant. Thyroid lesions without pathologic correlation were classified as either benign or indeterminate based on US evaluation. Lesions that were cystic, predominantly cystic, solid and cystic with features consistent with a colloid nodule, or stable for at least 1 year were considered benign. Fisher exact tests, Spearman correlations, and logistic regression models were used to evaluate the associations between CT and US characteristics and CT predictors of benignity or malignancy. Multivariable logistic regression analyses were performed to examine the joint associations between the CT features and sonographically assessed benignity. Of 259 patients, 168 (64.9%) patients had incidentally detected thyroid lesions, 49 (18.9%) patients had palpable/symptomatic lesions, and 116 (44.8%) patients underwent surgical resection and/or FNA biopsy. The malignancy prevalence was 1.8% (3 of 168) for incidental thyroid lesions and 10.2% (5 of 49) for palpable/symptomatic lesions. The malignancy prevalence of incidental lesions initially detected on CT was 1.6% (2 of 125). Of the 143 patients without pathologic data, 58 (40.6%) were classified as benign and 85 (59.4%) were categorized as indeterminate based on US evaluation. Statistically significant associations were found between CT and US with regard to lesion number, dominant lesion size, lesion consistency/composition, and associated calcifications. No CT characteristics of thyroid lesions predicted malignancy. However, there were statistically significant associations on multivariate analysis between indeterminate/benign nodules and CT characteristics of smaller lesion size, lower mean attenuation, and homogeneous composition. Recommending sonographic evaluation of all incidentally detected thyroid lesions is likely not the appropriate strategy, given the high prevalence of thyroid incidentalomas, low probability of malignancy, and cost effectiveness of workup. Small, homogeneous, low-attenuation lesions have a high probability of being benign.

Application of BRAF, NRAS, KRAS mutations as markers for the detection of papillary thyroid cancer from FNAB specimens by pyrosequencing analysis

BRAFV600E, the most common BRAF gene mutation, is detected in approximately 50% of sporadic papillary thyroid carcinoma (PTC) and may be associated with triggering tumorigenesis of PTC. The aim of our study was to discover additional mutations to increase the diagnostic performance of molecular tests in screening for thyroid cancer from fine needle aspiration biopsy (FNAB) specimens. DNA was extracted from 120 freshly obtained FNAB specimens selected according to cytopathology grades of the Bethesda system. A conventional BRAF V600E test was carried out with real-time PCR, and further mutation screening for BRAF mutations in codons 464, 466, 469, NRAS and KRAS codons 12/13 and 61 was done by pyrosequencing. Histopathology reports were reviewed for those who underwent thyroidectomy (n=83). The real-time PCR method detected 45 BRAF V600E- positive cases whereas pyrosequencing detected 30 cases. Additional BRAF (n=4), NRAS (n=11) and KRAS (n=3) mutations were detected in 17 cases (one overlapping BRAF and NRAS mutation). Among 11 NRAS-mutated cases, eight were confirmed as PTC and one as FVPTC on histopathology reports. Five PTC-confirmed cases with BRAF V600E mutation showed additional mutations, all of which were NRAS mutations. Despite the higher sensitivity of real-time PCR for detecting BRAFV600E mutations, pyrosequencing easily detected additional point mutations. NRAS mutations were the most prevalently identified additional mutations and were highly associated with malignancy. In conclusion, our findings demonstrate that additional mutations identified by pyrosequencing may help in the pre-operative process in determining the possibility of malignancy and further studies on the occurrence of simultaneous mutations of BRAF, KRAS and NRAS may be warranted.

Chromosome banding analysis of cells from fine-needle aspiration biopsy samples from soft tissue and bone tumors: is it clinically meaningful?

Morphologic evaluation of samples from fine-needle aspiration (FNA) and core needle (CN) biopsies is an important part of the pretreatment diagnosis of bone and soft tissue tumors. Because most such tumors have characteristic, sometimes even specific, chromosomal rearrangements, ancillary genetic analyses could provide important diagnostic information. Whereas directed analyses, such as fluorescence in situ hybridization or reverse transcriptase−polymerase chain reaction, for specific genetic aberrations are well suited for the relatively small cell numbers obtained with FNA biopsies, the possibility to obtain tumor karyotypes after cell culturing has been less well studied. In the present study, karyotypes from 114 FNA biopsy samples were compared to those in corresponding surgical tumor samples; in addition, results on 31 CN samples and their corresponding tumor samples were available. Of the 138 surgical tumor samples, 88 (64%) showed clonal acquired chromosome aberrations, 42 (30%) displayed a normal karyotype, and 8 (6%) did not yield any karyotype as a result of infection or poor cell growth. The corresponding figures for the 114 FNA samples were 27 (24%), 28 (25%), and 59 (52%), and for the 31 CN samples 15 (48%), 10 (32%), and 6 (19%). The relatively low success rate, with the possible exception of primitive round cell/Ewing sarcomas (abnormal karyotype in 6 of 11 FNA samples), strongly indicates that it is not meaningful to attempt cell culturing and chromosome banding analysis on FNA biopsy sample cells in patients with suspected bone or soft tissue tumors. The use of ancillary techniques such as fluorescence in situ hybridization might improve the diagnostic value from FNA. Our preliminary data suggest that if a pretreatment karyotype is wanted, the cytogenetic analysis should be made on cells from CN samples, close to half of which showed an aberrant karyotype.

Molecular analyses of thyroid tumors for diagnosis of malignancy on fine-needle aspiration biopsies and for prognosis of invasiveness on surgical specimens

High throughput genetic and genomic analyses have allowed the identification of series of genes exhibiting either distinct expression profiles or a particular mutational status in the different types or subtypes of thyroid tumors. The use of molecular data to improve the preoperative diagnosis of thyroid cancer on materiel from fine-needle aspiration biopsy (FNAB) is in the course of validation by numerous teams throughout the world. We have proposed a molecular test based on the expression level of a series of 19 genes, capable of discriminating malignant from benign tumors [15]. A prospective study aiming at the clinical validation of the molecular test has been performed on a cohort of 730 patients with a thyroid nodule. In patients subjected to tumor resection (≈ 220), the preoperative molecular diagnosis (generated on FNAB material from analyses of the expression level of the 19 genes) was compared to the postoperative diagnosis given by the pathologist (used as reference). Treatment and follow-up of the serious forms of thyroid cancer should benefit by the early identification of tumors with a metastatic potential using molecular characteristics differentiating invasive and non-invasive thyroid carcinomas. We have performed genetic and genomic analyses on a series of 200 papillary thyroid carcinomas (non-invasive or NI-PTC, 50%; invasive or I-PTC, 50%). BRAF V600E mutation or/and RET/PTC gene rearrangement have been detected in less than 25% of NI-PTC but in more than 75% of I-PTC. Pan-genomic analyses (Agilent microarray) revealed that 1373 genes are differentially expressed (fold change greater than 2) in NI-PTC as compared to I-PTC samples. The majority of genes (≈ 1200) are overexpressed in I-PTC. Data related to the two domains: diagnosis and prognosis of thyroid cancer will be presented at 2011 International H.P. KLOTZ conference on Clinical Endocrinology.

Thyroid tumor marker genomics and proteomics: Diagnostic and clinical implications

Two systems biology concepts, genomics and proteomics, are highlighted in this review. These techniques are implemented to optimize the use of thyroid tumor markers (TTM). Tissue microarray studies can produce genetic maps and proteomics, patterns of protein expression of TTM derived from preoperative biopsies and specimens. For instance, papillary and medullary thyroid cancers harbor RAS, RET, and BRAF genetic mutations. Follicular thyroid cancers harbor translocations and fusions of certain genes (PAX 8 and PPAR-gamma). Proteomic analysis from various tissue sources can provide useful information regarding the overall state of a thyroid cancer cell. Understanding the molecular events related to these genetic and protein alterations can potentially clarify thyroid cancer pathogenesis and guide appropriate molecular targeted therapies. However, despite the realization that these emerging technologies hold great promise, there are still significant obstacles to the routine use of TTM. These include equivocal thyroid nodule tissue morphologic interpretations, inadequate standardization of methods, and monetary costs. Interpretative shortcomings are frequently due to the relative scarcity of cellular material from fine-needle aspiration biopsy (FNAB) specimens. This can be rectified with large needle aspiration biopsy (LNAB) techniques and is exemplified by the favorable performance of galectin-3 determinations on LNAB specimens.

Cytologic evaluation of image-guided fine needle aspiration biopsies via robotic microscopy: A validation study

Background: This study carried out was to assess the feasibility of using robotic microscopy (RM) for cytologic evaluation of direct smears from fine needle aspiration biopsy (FNAB). Methods: Three board-certified cytopathologists reviewed representative direct smears from 40 image-guided FNABs using RM and subsequently re-reviewed the same smears using conventional microscopy. Adequacy of the smears and cytologic diagnosis, as determined using the two approaches, were compared for each individual cytopathologist (intraobserver) and between the three cytopathologists (interobserver). The intraobserver and interobserver discrepancies were analyzed and discussed in a follow-up consensus conference. Results: For assessment of adequacy, there were high concordance rates (intraobserver: 92.5-97.5%; interobserver: 90-92.5%), with a few discrepancies involving distinctions between suboptimal and satisfactory smears. Analysis of diagnostic interpretations showed correct classification of 92.5-95% (intraobserver) or 90-92.5% (interobserver) of benign and malignant cases combined, with the discrepancies being between benign and atypical cells in the benign group, and between suspicious and malignant in the malignant group. Within the malignant group, 94% of cases were accurately subclassified via RM. The quality of images viewed by using RM was rated adequate (fair or good) for 95% of the slides. Conclusions: The results demonstrate that cytologic evaluation of direct smears from FNABs using RM is feasible. Problems encountered included the longer times needed to evaluate cases with thick, bloody smears and/or low numbers of diagnostic cells, and difficulties in recognizing neuroendocrine differentiation and mimics of hepatocellular carcinoma.

Utilidad de la detección de tiroglobulina en el aspirado de punción ganglionar cervical en el seguimiento de pacientes con cancer papilar de tiroides

During the detection of neck recurrence in patients with Papillary Thyroid Carcinoma (PTC), sometimes it is difficult to distinguish metastatic from inflammatory neck lymph nodes. The measurement of serum thyroglobulin (sTg) under thyroid hormone suppression therapy the presence of serum thyroglobulin antibodies (sAbTg), the diagnostic whole body scan and cytology can give false negative results. Measurement of thyroglobulin in the washout fluid from fine-needle aspiration biopsy (FNAB) of suspicious neck lymph nodes could improve the diagnostic accuracy.To evaluate the usefulness of detecting Tg in lymph nodes (LTg) suspicious by ultrasonography (US) and compare it to cytology.Between the years 2004 and 2007 we prospectively studied 30 patients with PTC and cervical US findings of suspicious recurrence. LTg was assayed in US guided FNAB used for cytology.Sixteen out of 30 patients underwent surgery using as selective criteria an LTg higher than sTg or a positive cytology. Surgery confirmed the presence of metastasis in all 15 patients with positive LTg (8 with positive cytology) and in 1 patient with negative LTg and positive cytology (a case with undifferentiated thyroid cancer). The sensitivity was 93.7% for LTg and 56.2% for cytology. We identified by LTg 3 of 6 patients with undetectable sTg and positive sAbTg.The presence of LTg showed a higher sensitivity than cytology for the detection of cervical lymph node metastasis. This method is useful even in the presence of sAbTg.

Punção aspirativa nos tumores das glândulas salivares: especificidade e sensibilidade

Neoplasms of salivary glands represent almost 3% of all head and neck tumors. Proper surgical treatment depends upon accurate histological findings, especially in the case of malignant lesions. As such, knowledge of correct cytological findings prior to surgery is important for therapeutic planning. This is not easily established since it is usually based only on the patients' clinical history and imaging exams To evaluate results obtained from fine needle aspiration biopsy (FNAB), comparing them to the histological findings of the respective surgical specimens and analyzing the sensitivity, specificity and accuracy of this method in relation to the diagnosis of malignancy or benignancy. Retrospective study with the medical records of 73 patients who had salivary gland neoplasms and were submitted to FNAB and surgical treatment. Data disclosed that values of sensitivity were 87.9 % and specificity 85.7 % for diagnosis of benign tumors. For malignant tumors 42.5 % of sensitivity and 98.3 of specificity, were observed. Overall values of accuracy were, respectively, 85.7 % and 87.8 % for positive and negative predictive values in the diagnosis of malignancy by FNAB. It was shown that FNAB, as supplementary diagnostic method, can be useful for preoperative evaluation and surgical planning especially for malignant neoplasms.

The role of ThinPrep cytology in the evaluation of estrogen and progesterone receptor content of breast tumors

The objective of the current study was to analyze the potential value of immunocytochemical analysis on ThinPrep (TP)-processed smears, from fine needle aspiration (FNA) biopsies, of breast tumors for the determination of ER and PR content as compared with the immunohistochemical analysis performed on paraffin-embedded breast tumor specimens. Percutaneous FNA biopsy of focal breast lesions in 119 female adult patients during a 31-month period was performed. Subsequently, these patients underwent surgical resection of the tumors. ER and PR status of the tumors was determined by immunocytochemical analysis on TP-processed smears and by immunohistochemical studies in paraffin-embedded sections. With the use of TP technique adequate material was observed in all cases. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and overall accuracy (OA) of the TP technique for the ER were 98.95%, 100%, 100%, 95.84% and 99.15% respectively. In addition, sensitivity, specificity, PPV, NPV and OA of the TP technique for the PR were 100%, 87.5%, 95.60%, 100% and 96.64%, respectively. ER and PR status can be evaluated in FNA material from breast carcinomas by using the TP technique. Sample collection and storage is simple and permits the assortment of the FNA sample for both morphologic diagnosis and ancillary studies. The accuracy of TP technique in the detection of ER and PR content is comparable to those of the histological evaluation, and could be of paramount importance for the preoperative planning of treatment.

Isolation of Thyroid Cells Obtained by Fine-Needle Aspiration Biopsy

Usually thyroid cells isolated from tissue obtained by surgery or thyroid cell lines are used to investigate the pathogenesis of autoimmune thyroid diseases. Isolation and cultivation of thyrocytes from fine-needle aspiration biopsy (FNAB) has not yet been published. The aim of this study was to isolate and cultivate thyrocytes from samples of FNAB. FNAB samples were obtained from nine adults and nine children with Hashimoto's thyroiditis (HT). The aspiration material was filtered resulting in small samples of tissue on the surface of the filter membrane. These tissue fragments were digested by collagenase I and dispase II. The yielding cells were cultivated for 3 weeks in Ham's F12 Kaighn's Modification medium in presence of 1 mU/mL bovine thyrotropin (TSH), 10 microg/mL human insulin, 6 microg/mL transferrin, and 10(-8) M hydrocortisone. Finally, isolated thyroid cells were characterized by determination of gene expression of thyrotropin receptor (TSHR), thyroperoxidase (TPO), and thyroglobulin (Tg) using a nested reverse transcriptase-polymerase chain reaction (RT-PCR). Thyroid cells obtained by FNAB can be maintained over a time period of approximately 3 weeks. Depending on the sample size a final number of 1000-14,000 cells was gained per FNAB. In addition, all cells isolated by the described method expressed TPO mRNA. TSHR mRNA was found in 4 samples, whereas 15 samples were Tg mRNA-positive. There were no differences with respect to the expression TSHR and TPO mRNA between samples from adults and children. The isolation and cultivation of thyroid cells obtained by FNAB has been established. In contrast to surgical specimen, this technique provides an easy access to thyrocytes derived from individual patients allowing repeated sampling to investigate the time progression of the chronic disease or the effect of treatment over time.

CD56 expression of neuroendocrine neoplasms on immunophenotyping by flow cytometry: a novel diagnostic approach to fine-needle aspiration biopsy.

CD56 antigen or NCAM (neural cell adhesion molecule) has an established role in the diagnosis of non-Hodgkin lymphoma (NHL)-natural killer cell type and other hematologic malignancies. Therefore, it is included routinely in the panel of antibodies for flow cytometric (FC) analysis of suspected lymphomatous tissue specimens obtained from fine-needle aspiration biopsy (FNAB). The authors evaluated the role of CD56 expression on FC of neuroendocrine (NE) tumors. An initial diagnosis of NHL was suspected based on an on-site FNAB evaluation. Ten FNABs were identified from the cytopathology files at The Johns Hopkins Hospital, Baltimore, MD (2000-2001). Flow cytometric analysis was negative for NHL but revealed a CD56-positive nonlymphoid cell population. An FNAB evaluation was performed on air-dried Diff-Quik-stained smears and FC analysis used a fixed panel of 12 antibodies (B-cell markers, T-cell markers, CD33, CD56, and CD71). Immunoperoxidase staining (IPOX) was performed on the cell block sections from four of the tissue specimens using epithelial and NE markers, CD56, desmin, and O13 antibodies. Sites of FNAB included the lung (five cases), liver (one case), lymph node (three cases), and peritoneum (one case). Only one patient had a history of cancer at the time of FNAB. All cytologic diagnoses were confirmed by histopathologic follow-up on resection or biopsy or both. Diagnoses included small cell carcinoma (eight cases), Merkel cell carcinoma (one case), and primitive neuroectodermal tumor/Ewing sarcoma (one case). All tissue specimens that underwent IPOX stained strongly with NE markers, with one tissue section staining only with O13. CD56 expression by FC in the presence of negative immunostaining with lymphoid markers represented a unique yet highly specific method for the diagnosis of NE tumors by FNAB. This procedure eliminated the need for further IPOX studies on the already limited cytologic sample and provided a timely and accurate diagnosis.

Heterogeneity of the thyroglobulin epitopes associated with circulating thyroid hormone autoantibodies in hashimoto's thyroiditis and non-autoimmune thyroid diseases.

We previously implicated TG leakage from fine-needle aspiration biopsy (FNAB) as responsible for circulating thyroid hormone autoantibodies (THAb). However, THAb were not always associated with TGAb. In the literature these negative findings have been interpreted against a role of TG as the antigen for THAb. To evaluate the TGAb status more fully and to gain information on TG epitopes involved in THAb development, we measured: 1) TGAb with an independent hemagglutination assay (HA), and 2) epitope specificity in a competitive ELISA using 2 monoclonal Abs (mAb) against TG: mAb 42C3 and mAb 134C2. MAb 42C3 recognizes a cross-reactive iodinated epitope, whereas 134C2 is specific for human TG. We tested 12 Hashimoto's thyroiditis (HT) and 35 non-HT patients sampled prior to, 1 and 3 months after FNAB. We found that, irrespective of thyroid disease or post-FNAB THAb status, certain patients previously classified as TGAb negative by IRMA tested TGAb positive by HA or by competition ELISA and vice versa. A post FNAB positive response to the 42C3 iodinated epitope in only one THAb IgM-T4+ve HT and a few THAb negative non-HT patients was observed. Furthermore, we observed that the 3 non-HT patients who expressed IgM-T3 THAb failed to bind either TG-mAb epitope. We conclude that a single TGAb assay is not sufficient to define the TGAb status, which can be achieved reliably only by using multiple TGAb assays. In addition, the TG-iodinated epitope recognized by 42C3 is not a major epitope in post-FNAB THAb, and the T3-epitope involved in THAb remains distinct from the mAb epitopes. In light of recent data in the literature, we further suggest that the responsible epitopes are more likely to be expressed in leaked TG fragments, rather than leaked intact TG.

Optimal Recovery of DNA for Polymerase Chain Reaction–Based Assays from Fine Needle Aspirates

To assess the ideal preparation for molecular techniques applied to cytologic specimens using polymerase chain reaction (PCR) on different types of cytologic preparations with specimens obtained from fine needle aspiration biopsy (FNAB). Besides conventional cytologic examination, PCR was performed on 30 consecutive cases of FNAB to analyze the beta-actin gene in four types of cytologic preparations, including fresh samples, previously stained Papanicolaou and Diff-Quik routine smears and cell blocks. Cellularity and cytologic findings were correlated with PCR results. The beta-actin gene was successfully amplified from all cases studied. Cellularity of the samples correlated directly with PCR results except for one case of metastatic melanoma with intense pigment retention. All specimens showed equivalent results as compared to fresh samples. Generally cell blocks were less cellular and consequently less effective. All conventional cytologic preparations tested were suitable for PCR-based assays when cellularity was adequate. Alcohol-fixed and air-dried smears can be successfully used for PCR studies, providing rapid and simple specimens for molecular studies, which can add important information concerning diagnosis, prognosis and management.

Immunostaining for Met/HGF receptor may be useful to identify malignancies in thyroid lesions classified suspicious at fine-needle aspiration biopsy.

The receptor for hepatocyte growth factor (Met) is not expressed in the normal thyroid but it is overexpressed in most thyroid carcinomas. We evaluated whether Met immunostaining of cytological smears from fine-needle aspiration biopsy (FNAB) may be useful for the preoperative diagnosis of thyroid cancer. Notably, routine cytological examination often fails to distinguish well-differentiated follicular carcinomas and a proportion of papillary carcinomas (low-grade papillary carcinomas and papillary carcinomas follicular variant [FVPTC]) from benign lesions: all these lesions are usually classified as suspicious. We examined 80 thyroid lesions diagnosed as suspicious at cytology that had subsequently undergone surgery. The histologic diagnosis had been: papillary carcinomas (n = 14), FVPTC (n = 11), follicular carcinomas (n = 25), atypical follicular adenomas (n = 5), follicular adenomas (n = 20), and nodular goiters (n = 5). We also studied typical papillary carcinomas (n = 30) and nodular goiters (n = 10), all correctly diagnosed at cytology. In lesions classified suspicious at routine cytology, Met immunostaining was positive in 12 of 14 (85.7%) papillary carcinomas, 8 of 11 (72.7%) FVPTC, 7 of 25 (28%) follicular carcinomas, and 5 of 5 atypical adenomas. In contrast, none of the 25 lesions cytologically suspicious but benign at histology were positive. These data suggest that Met immunostaining of suspicious cytological smears are useful for identifying malignant lesions, especially those with a papillary histotype.

Rhabdomyosarcoma

To identify morphologic characteristics and architectural patterns of rhabdomyosarcoma (RMS), to attempt a subclassification from fine needle aspiration biopsy (FNAB) smears and to point out some differential diagnostic problems. We reviewed all positive cytologic material from 53 patients with RMS whose diagnoses were histologically and/or immunocytochemically confirmed. We analyzed several morphologic features and identified architectural patterns of smears. Among alveolar RMS, we identified two major architectural patterns: one containing completely dissociated cells and one containing many chance formations. Among the embryonal type, the predominant architectural pattern contained large tissue fragments with abundant eosinophilic material and various numbers of dissociated cells. The pattern of only dissociated cells was similar to the one seen in the alveolar type. The relative proportion of poorly to better and well-differentiated rhabdomyoblasts varied in both types and in all patterns. RMS exhibits a variety of morphologic pictures regarding cellular morphology and architectural patterns, even within the same histologic subtype. Therefore, a reliable subclassification into alveolar and embryonal RMS cannot be made from FNAB smears. The embryonal type can be suggested in cases containing large tissue fragments with abundant eosinophilic material and small, tightly packed cells with oval nuclei. However, all cases suspected to be RMS must always be confirmed immunocytochemically since they could be confused with some benign and malignant tumors with similar morphology.

DNA flow cytometry of non-Hodgkin's lymphomas: correlation with cytologic grade and clinical relapse.

In this prospective study, we correlated cytological grading and clinical follow-up of non-Hodgkin's lymphomas (NHL) with DNA flow cytometry (FCM) data from fine-needle aspiration biopsy (FNAB) material. FNAB was performed from 34 successive cases of NHL, and the aspirated material was analyzed by DNA FCM. Cytological subtyping and grading were done by modified Working Formulation. Cases were followed up for 2.5 yr, and cytological grading, clinical follow-up, and DNA FCM data were correlated. There were 8 cases of low, 14 intermediate, and 12 high-grade NHL. None of the cases of low-grade NHL showed DNA aneuploidy. Of the 8 DNA aneuploid cases, 5 were of intermediate and 3 were of high-grade NHL. Mean growth fraction (GF) of low-grade, intermediate-grade, and high-grade NHL was 6.1, 9.4, and 19.4, respectively. DNA aneuploidy was statistically significant (P < 0.05) between low- vs. intermediate- and high-grade NHL. Growth fraction was also statistically significant between low- vs. high-grade NHL. Six cases showed clinical recurrence, and one case died within 6 mo of diagnosis, due to widespread involvement of NHL. DNA aneuploidy and mean GF of recurrent and nonrecurrent cases were statistically significant (P < 0.05) irrespective of grading of NHL. DNA aneuploidy and GF are not well-correlated with morphological grading (by the Working Formulation). However, these two criteria are important for assessment of early clinical relapse of NHL.