PubMed · 15511469
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Paul H Schipper, Francis C Nichols. 2004. Invited commentary.. https://doi.org/10.1016/j.athoracsur.2004.06.073
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We present a method based on spatially resolved electron energy-loss spectroscopy in the cryo-STEM to map the spatial distribution of water in frozen-hydrated polymers. The spatial resolution is limited by the dose constraints imposed by radiation damage, and to stay within these constraints, the use of fine electron-probe sizes comes at the cost of reduced counts in the energy-loss spectra. Thus, at the resolution limit, the detection of isolated water-rich pixels or the identification of minor variations in water content across the specimen is complicated because one must distinguish significant fluctuations from noise. Here we develop a criterion with which to guide such a distinction. We characterize the intrinsic noise associated with spectral measurements under given illumination and acquisition conditions. We then use that noise in combination with scatter diagrams to threshold spectrum images and objectively identify statistically significant compositional fluctuations. We illustrate these ideas using a simulated spectrum dataset for a hypothetical blend of hydrophilic and hydrophobic homopolymers. We show that while a direct inspection of the water map may not allow any meaningful conclusions to be drawn, after applying the thresholding approach we can clearly identify the regions of the specimen that are rich in water. We also experimentally study a model blend system comprised of hydrophilic poly(vinyl pyrrolidone) (PVP) dispersed in a hydrophobic matrix of poly(styrene) (PS). By MLS fitting using damaged and undamaged PVP reference spectra, we determine that the critical dose characteristic of dry PVP is approximately 8000 e/nm2 using 200 keV incident electrons. Irradiating frozen-hydrated PVP gives rise to noticeable hydrogen evolution at doses of approximately 1500 e/nm2. To stay within this constraint we use doses of 400 e/nm2 and a pixel spacing in the spectrum imaging of 100 nm. In order to quantitatively map the water, PVP, and PS compositions, we measure their total inelastic scattering cross-sections. Direct inspection of the composition maps reveals the presence of large water-rich domains of the order of approximately 1 microm and the scatter-diagram thresholding approach identifies small water-rich domains one pixel in size.
OBJECTIVE: Intraoperative identification of parathyroid tissue is crucial during parathyroid surgery. Frozen section is the most common tool, but is time-consuming and expensive. Scrape cytology is a modification of imprint cytology that provides rapid and cheap intraoperative identification of parathyroid tissue, but its reliability remains controversial. We assessed the sensitivity and specificity of scrape cytology in the intraoperative identification of parathyroid tissue. METHODS: Scrape cytology samples from parathyroid tissue (n=29), adipose tissue (n=3) and thyroid tissue (n=2) were prepared intraoperatively using May-Grünwald-Giemsa and Papanicolaou staining and subsequently sent for routine frozen section and standard paraffin section. A single pathologist reading the scrape cytology was blinded to the results of the frozen and standard paraffin sections. RESULTS: Scrape cytology identified 25 of 29 parathyroid tissues, all three adipose tissues and one of two thyroid tissues. The remaining samples, four parathyroid tissues and one thyroid tissue, were not identified due to insufficient sample for diagnosis. The result translated to give a diagnostic accuracy of 88.2%, sensitivity of 86%, and specificity of 100%. All tissues were accurately identified by frozen section. CONCLUSION: Scrape cytology is a rapid, economical test with acceptable sensitivity and high specificity. It can be used as an adjunct to frozen section and may be used as a tool in helping surgeons to identify parathyroid tissue.
BACKGROUND: Immunohistochemistry has revolutionized the use of frozen sections for the removal of melanoma. OBJECTIVE: Two studies were performed to address the best and fastest way to stain melanocytes in frozen tissue sections. MATERIALS AND METHODS: Fifty-six standing cones were taken from sun-damaged white skin and stained with combinations of anti-S-100, anti-HMB-45, anti-Melan-A, and azure B with appropriate controls. RESULTS: Anti-Melan-A was the best immunohistochemical stain. Azure B stains melanin granules metachromatically green and allows differentiation of melanocytes and melanophages. Ultrarapid staining worked as well as rapid staining. CONCLUSION: Dako's EnVision ultrarapid staining protocol and azure B counterstain represent a dramatic improvement and should be considered the easily obtainable histologic standard when excising cutaneous melanoma.