Serial sections
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- MeSH
- diencefalon anatomie a histologie MeSH
- experimenty na zvířatech MeSH
- histologické techniky * metody využití MeSH
- kočky MeSH
- mezencefalon anatomie a histologie MeSH
- mozek * anatomie a histologie fyziologie MeSH
- výzkumné techniky MeSH
- zmrazené řezy * metody využití MeSH
- zvířata MeSH
- Check Tag
- kočky MeSH
- zvířata MeSH
- Publikační typ
- fotografie MeSH
2nd compl. rev. and expanded ed. VIII, 213 s. : il.
Three-dimensional (3-D) reconstruction from microscopic images represents a useful tool for the study of biological structures in embryology and developmental biology. However, it is usually necessary to cope with many difficulties connected with the preparation of specimens. In order to minimize mutual displacement of structures in successive sections, the applicability of non-deparaffinized tissue sections for 3-D reconstruction was tested. Chicken embryos were fixed and stained in toto with eosin and then embedded in paraffin. About 30-mum-thick non-deparaffinized serial sections were used for obtaining initial data for 3-D reconstruction of larger stacks of embryonic bodies using either fluorescence or confocal microscope. The same sections served for both collecting optical serial sections of mesonephros as source images for its 3-D reconstruction, and immunohistochemical detection of fibronectin, laminin and vimentin. It was found that sections with retained paraffin preserve the mutual spatial relationships of tissue components as well as provide an excellent differentiation of structure. It makes the process of 3-D reconstruction easier. The localization of the products of immunohistochemical reactions demonstrated the co-localization of fibronectin and laminin in basal laminas and the presence of vimentin in glomeruli and mesenchymal tissue. The use of non-deparaffinized sections represents a less time consuming and more effective alternative to thin histological sections for the purpose of 3-D reconstruction, and enables further application of material.
- MeSH
- barvení a značení MeSH
- biologické markery metabolismus MeSH
- fibronektiny metabolismus MeSH
- fluorescenční mikroskopie MeSH
- imunoenzymatické techniky MeSH
- konfokální mikroskopie MeSH
- kuřecí embryo MeSH
- laminin metabolismus MeSH
- mezonefros cytologie embryologie metabolismus MeSH
- mikrotomie MeSH
- počítačové zpracování obrazu metody MeSH
- vimentin metabolismus MeSH
- zalévání tkání do parafínu MeSH
- zvířata MeSH
- Check Tag
- kuřecí embryo MeSH
- zvířata MeSH
Jsou uváděny výsledky histologického vyšetření sentinelových lymfatických uzlin (SLN) u karcinomu prsu metodou rychlé peroperační biopsie ze zmrazených řezů. Popisujeme první zkušenosti ze souboru 77 pacientek, řešených v období březen 2005 – červenec 2006 záchovným resekčním výkonem s peroperační biopsií axilárních sentinelových uzlin a v případech negativního nálezu pak následným zpracováním bloků v sériových řezech za použití imunohistochemie. Našim cílem bylo ověřit, jaké množství metastáz zachytíme v průběhu samotné peroperační biopsie (senzitivita) a jaký je poměr negativních nálezů s pozitivitou v následném sériovém zpracování k celkovému množství peroperačně negativních nálezů (falešná negativita). Získané výsledky jsou porovnány s recentně publikovanými pracemi. Krátce je diskutována náročnost a možnosti zvolené metody. V souboru 77 pacientek s karcinomem prsu bylo vyšetřeno celkem 193 SLN, medián 2,5 (rozmezí 1–7 uzlin). Po kompletním vyšetření byly sentinelové uzliny negativní ve 45 případech (58,4 %). Ve 32 případech bylo zjištěno metastatické postižení SLN (41,6 %), z toho peroperačně jsme zjistili metastázu ve 24 případech, to je 75%senzitivita. Specificita je přitom 100%. V 8 dalších případech byly zjištěny metastázy až v následném sériovém zpracování bez nebo s použitím imunohistochemie – falešná negativita 15,1%. Ve všech případech se jednalo o mikrometastázy, nebo izolované nádorové buňky (ITC) a buněčné klastry.
Results of histological examinations of sentinel lymphatic nodes (SLN) obtained by rapid peroperative biopsy (RPB) are presented. Our first experience with 77 patients undergoing localized excision of axillary sentinel nodes is reported. Negative nodes were subsequently examined by means of immunohistochemistry of serial sectioned blocks. The aim of the study was to verify the percentage of identified metastases, and thus define the reliability of the RPB. At the same time we tried to determine the ratio of the negative biopsies of SLN, which were found positive subsequent to immunohistochemistry examination of serial sections, to the total amount of peroperative negative findings or a “false negativity”. Our results were compared with those recently published. Particular demands and possibilities of the method used are briefly discussed. In the group of 77 patients with breast cancer, the total number of 193 SLN were examined (average 2.5, in a patient ranging from 1 to 7). Out of all examined SLN, 45 patients (58.4%) were negative. Metastases were identified in 32 patients (41.6%). By rapid preoperative biopsy alone, metastases were found in 24%, which represents 75% sensitivity. The specificity was 100%. The following examination of serial sectioned specimens with or without immunohistochemistry showed 8 more patients with metastases, which represents the false negativity of 15.1%. The metastases found in all 8 patients were small micrometastases, isolated tumour cells or clusters of cells.
A set of methods leading to volume reconstruction of biological specimens larger than the field of view of a confocal laser scanning microscope (CLSM) is presented. Large tissue specimens are cut into thin physical slices and volume data sets are captured from all studied physical slices by CLSM. Overlapping spatial tiles of the same physical slice are stitched in horizontal direction. Image volumes of successive physical slices are linked in axial direction by applying an elastic registration algorithm to compensate for deformations because of cutting the specimen. We present a method enabling us to keep true object morphology using a priori information about the shape and size of the specimen, available from images of the cutting planes captured by a USB light microscope immediately before cutting the specimen by a microtome. The errors introduced by elastic registration are evaluated using a stereological point counting method and the Procrustes distance. Finally, the images are enhanced to compensate for the effect of the light attenuation with depth and visualized by a hardware accelerated volume rendering. Algorithmic steps of the reconstruction, namely elastic registration, object morphology preservation, image enhancement, and volume visualization, are implemented in a new Rapid3D software package. Because confocal microscopes get more and more frequently used in scientific laboratories, the described volume reconstruction may become an easy-to-apply tool to study large biological objects, tissues, and organs in histology, embryology, evolution biology, and developmental biology. In this work, we demonstrate the reconstruction using a postcranial part of a 17-day-old laboratory Wistar rat embryo. (c) 2008 Wiley-Liss, Inc.
- MeSH
- algoritmy MeSH
- embryo savčí cytologie MeSH
- financování organizované MeSH
- konfokální mikroskopie metody MeSH
- krysa rodu rattus MeSH
- mikrotomie metody MeSH
- počítačové zpracování obrazu metody MeSH
- pružnost MeSH
- zobrazování trojrozměrné metody MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
