Is there an indication for frozen section examination of the ureteral margins during cystectomy for transitional cell carcinoma of the bladder?

PURPOSE: We evaluated the incidence of pathological findings of the ureter at cystectomy for transitional cell carcinoma of the bladder and assessed the usefulness of intraoperative frozen section examination of the ureter. MATERIALS AND METHODS: Histopathological findings of ureteral frozen section examination were compared to the corresponding permanent sections and the diagnostic accuracy of frozen section examination was evaluated. These segments were then compared to the more proximal ureteral segments resected at the level where they cross over the common iliac arteries. The histopathological findings of the ureteral segments were then correlated for upper urinary tract recurrence and overall survival. RESULTS: Transitional cell carcinoma or carcinoma in situ was found on frozen section examination of the distal ureter in 39 of 805 patients (4.8%) and on permanent sections in 29 (3.6%). In 755 patients the false-negative rate of frozen section examination of the ureters was 0.8%. Of the patients with carcinoma in situ diagnosed on the first frozen section examination 80% also had carcinoma in situ in the bladder. Transitional cell carcinoma or carcinoma in situ in the most proximally resected ureteral segments was found in 1.2% of patients. After radical cystectomy there was tumor recurrence in the upper urinary tract in 3% of patients with negative ureteral frozen section examination and in 17% with carcinoma in situ on frozen section examination. CONCLUSIONS: Routine frozen section examination of the ureters at radical cystectomy is only recommended for patients with carcinoma in situ of the bladder, provided the ureters are resected where they cross the common iliac arteries.

Parotid tumors: fine-needle aspiration and/or frozen section

OBJECTIVE: The purpose of this study was to analyze and compare the value of fine-needle aspiration cytology (FNAC) and frozen section (FS) analysis in the assessment of parotid gland tumors. STUDY DESIGN: Chart review and cross-sectional analysis. SUBJECTS AND METHODS: FNAC and FS analysis of 110 parotid tumors, 68 malignancies and 42 benign tumors, were analyzed and compared with the final histopathologic diagnosis. RESULTS: The accuracy, sensitivity, and specificity of FNAC in detecting malignant tumors were 79 percent, 74 percent, and 88 percent, respectively. On FS analysis, the accuracy, sensitivity, and specificity in detecting malignant tumors were 94 percent, 93 percent, and 95 percent, respectively. The histologic tumor type was correctly diagnosed by FNAC and FS in 27 of 42 (64%) and 39 of 42 (93%) benign tumors, respectively, and in 24 of 68 (35%) and 49 of 68 (72%) malignant neoplasms, respectively. CONCLUSION: The current analysis showed a superiority of FS compared with FNAC regarding the diagnosis of malignancy and tumor typing. FNAC alone is not prone to determine the surgical management of parotid malignancies.

Absence of ectopic epithelial inclusions in 3,904 axillary lymph nodes examined in sentinel technique

Intraoperative examination of sentinel axillary lymph nodes can be done by imprint cytology, frozen section, or, most recently, by PCR-based amplification of a cytokeratin signal. Using this technique, benign epithelial inclusions, representing mammary tissue displaced along the milk line, will likely generate a positive PCR signal and lead to a false-positive diagnosis of metastatic disease. To better appreciate the incidence of ectopic epithelial inclusions in axillary lymph nodes, we have performed an autopsy study, examining on 100 μm step sections 3,904 lymph nodes obtained from 160 axillary dissections in 80 patients. The median number of lymph nodes per axilla was 23 (15, 6, and 1 in levels 1, 2, and 3, respectively). A total of 30,450 hematoxylin-eosin stained slides were examined, as well as 8,825 slides immunostained with pan-cytokeratin antibodies. Despite this meticulous work-up, not a single epithelial inclusion was found in this study, suggesting that the incidence of such inclusions is much lower than the assumed 5% reported in the literature.

A new tool based on two micromanipulators facilitates the handling of ultrathin cryosection ribbons

A close to native structure of bulk biological specimens can be imaged by cryo-electron microscopy of vitreous sections (CEMOVIS). In some cases structural information can be combined with X-ray data leading to atomic resolution in situ. However, CEMOVIS is not routinely used. The two critical steps consist of producing a frozen section ribbon of a few millimeters in length and transferring the ribbon onto an electron microscopy grid. During these steps, the first sections of the ribbon are wrapped around an eyelash (unwrapping is frequent). When a ribbon is sufficiently attached to the eyelash, the operator must guide the nascent ribbon. Steady hands are required. Shaking or overstretching may break the ribbon. In turn, the ribbon immediately wraps around itself or flies away and thereby becomes unusable. Micromanipulators for eyelashes and grids as well as ionizers to attach section ribbons to grids were proposed. The rate of successful ribbon collection, however, remained low for most operators. Here we present a setup composed of two micromanipulators. One of the micromanipulators guides an electrically conductive fiber to which the ribbon sticks with unprecedented efficiency in comparison to a not conductive eyelash. The second micromanipulator positions the grid beneath the newly formed section ribbon and with the help of an ionizer the ribbon is attached to the grid. Although manipulations are greatly facilitated, sectioning artifacts remain but the likelihood to investigate high quality sections is significantly increased due to the large number of sections that can be produced with the reported tool.

Ceramic foam plates: a new tool for processing fresh radical prostatectomy specimens

Procurement of fresh tissue of prostate cancer is critical for biobanking and generation of xenograft models as an important preclinical step towards new therapeutic strategies in advanced prostate cancer. However, handling of fresh radical prostatectomy specimens has been notoriously challenging given the distinctive physical properties of prostate tissue and the difficulty to identify cancer foci on gross examination. Here, we have developed a novel approach using ceramic foam plates for processing freshly cut whole mount sections from radical prostatectomy specimens without compromising further diagnostic assessment. Forty-nine radical prostatectomy specimens were processed and sectioned from the apex to the base in whole mount slices. Putative carcinoma foci were morphologically verified by frozen section analysis. The fresh whole mount slices were then laid between two ceramic foam plates and fixed overnight. To test tissue preservation after this procedure, formalin-fixed and paraffin-embedded whole mount sections were stained with hematoxylin and eosin (H&E) and analyzed by immunohistochemistry, fluorescence, and silver in situ hybridization (FISH and SISH, respectively). There were no morphological artifacts on H&E stained whole mount sections from slices that had been fixed between two plates of ceramic foam, and the histological architecture was fully retained. The quality of immunohistochemistry, FISH, and SISH was excellent. Fixing whole mount tissue slices between ceramic foam plates after frozen section examination is an excellent method for processing fresh radical prostatectomy specimens, allowing for a precise identification and collection of fresh tumor tissue without compromising further diagnostic analysis.

Immunohistochemical panel for the diagnosis of Hirschsprung's disease using antibodies to MAP2, calretinin, GLUT1 and S100.

AIMS The diagnosis of Hirschsprung's disease is currently based on the identification of aganglionosis and the presence of an increase in acetylcholinesterase-positive hypertrophic nerve fibres in the large bowel submucosa. However, acetylcholinesterase staining is laborious and requires a skilled technician. The aim of this study was to identify a method for diagnosing Hirschsprung's disease reliably using an immunohistochemical panel of recently proposed markers. METHODS AND RESULTS Sixty-nine specimens from 37 patients were evaluated. MAP2 and calretinin antibodies were shown to stain ganglia reliably in the submucosal and myenteric plexuses of normal tissue. By contrast, reduced staining of ganglia was observed in patients with Hirschsprung's disease. Staining for GLUT1 and S100 was used to evaluate the number and thickness of nerve fibres. Gain of GLUT1 and S100 expression was in contrast to the loss of calretinin and MAP2. Hypertrophic submucosal nerve fibres in Hirschsprung's disease develop a perineurium with a ring-like GLUT1 staining pattern similar in size and intensity to that observed in deeper subserosal tissue. CONCLUSIONS The diagnosis of Hirschsprung's disease using immunohistochemical panels could be as accurate as with conventional frozen section techniques. In particular, the use of a combination of markers for ganglia and hypertrophic nerve fibres highlighting a prominent perineurium in Hirschsprung's disease could be an alternative method.

Electron microscopy of high pressure frozen samples: bridging the gap between cellular ultrastructure and atomic resolution

Transmission electron microscopy has provided most of what is known about the ultrastructural organization of tissues, cells, and organelles. Due to tremendous advances in crystallography and magnetic resonance imaging, almost any protein can now be modeled at atomic resolution. To fully understand the workings of biological "nanomachines" it is necessary to obtain images of intact macromolecular assemblies in situ. Although the resolution power of electron microscopes is on the atomic scale, in biological samples artifacts introduced by aldehyde fixation, dehydration and staining, but also section thickness reduces it to some nanometers. Cryofixation by high pressure freezing circumvents many of the artifacts since it allows vitrifying biological samples of about 200 mum in thickness and immobilizes complex macromolecular assemblies in their native state in situ. To exploit the perfect structural preservation of frozen hydrated sections, sophisticated instruments are needed, e.g., high voltage electron microscopes equipped with precise goniometers that work at low temperature and digital cameras of high sensitivity and pixel number. With them, it is possible to generate high resolution tomograms, i.e., 3D views of subcellular structures. This review describes theory and applications of the high pressure cryofixation methodology and compares its results with those of conventional procedures. Moreover, recent findings will be discussed showing that molecular models of proteins can be fitted into depicted organellar ultrastructure of images of frozen hydrated sections. High pressure freezing of tissue is the base which may lead to precise models of macromolecular assemblies in situ, and thus to a better understanding of the function of complex cellular structures.