Registration parameter optimization for 3D tissue modeling from resected tumors cut into serial H&E slides
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Patients diagnosed with early stage (Stage I/II) Oral Cavity Cancer (OCC) are typically treated with surgery alone. Unfortunately, 25-37% of early stage OCC patients experience loco-regional tumor recurrence after receiving surgery. Currently, pathologists use the Histologic Risk Model (HRM), a clinically validated risk assessment tool to determine patient prognosis. In this study, we perform image registration on two cases of serially sectioned blocks of Hematoxylin and Eosin (H and E) stained OCC tissue sections. The goal of this work is to create an optimized registration procedure to reconstruct 3D tissue models, which can provide a pathologist with a realistic representation of the tissue architecture before surgical resection. Our project aims to extend the HRM to enhance prediction performance for patients at high risk of disease progression using computational pathology tools. In previous literature, others have explored image registration of histological slides and reconstructing 3D models with similar processes used. Our work is unique in that we are investigating in-depth the parameter space of an image registration algorithm to establish a registration procedure for any serial histological section. Each parameter set was sequentially perturbed to determine the best parameter set for registration, as evaluated through mutual information.Keywords:
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Reinke crystalloid (CR)s are rod or corn shaped structures present in the interstitial cells (Leydig cell) of human testis. Due to existing controversies and because of their importance in histological studies, we decided to reevaluate their staining behavior with various dyes. Earlier it has been shown that hematoxylin and eosin dyes do not stain CRs even though their protein nature remains undisputed. In the present study, sections of testicular tissues embedded in glycol-methacrylate (GMA) showed that hematoxylin binds to CRs non-specifically and eosin stains them specifically. The reasons for stainability and/or non stainability of CRs with hematoxylin, eosin and other dyes are discussed.
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Histopathology is often essential to establish an accurate diagnosis. Pathology laboratories are scarce in most Sub-Saharan Africa where dermatopathology is a developing field. In resource-poor countries, most specimens are analyzed only after hematoxylin and eosin staining. The availability of special stains is very limited and restricted to only few centers. The aim of this study is to analyze the extent of dermatopathological cases which can be adequately diagnosed after hematoxylin and eosin alone. Secondly, to investigate which cases required further special stains.All skin specimens submitted to two University Hospitals (Tanzania and Kenya) were included in this study. All specimens were first analyzed with hematoxylin and eosin and a diagnosis established when possible. All cases in which an accurate diagnosis after hematoxylin and eosin only was not possible, were registered and evaluated after further special stains.A total of 386 specimens were examined. A proper histopathologic diagnosis with hematoxylin and eosin alone was possible in 344 (89.1%) samples. In 45 (11.6%) cases, mostly skin infections, further special stains were necessary.A proper histopathologic diagnosis was possible after hematoxylin and eosin alone in almost 90% of the specimens submitted to the two laboratories in Sub-Saharan Africa.
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You have accessJournal of UrologyAdult urology1 Feb 2006Virtual Microscopy in Prostate Histopathology: Simultaneous Viewing of Biopsies Stained Sequentially With Hematoxylin and Eosin, and α-Methylacyl-Coenzyme A Racemase/p63 Immunohistochemistry Henrik O. Helin, Mikael E. Lundin, Mervi Laakso, Johan Lundin, Heikki J. Helin, and Jorma Isola Henrik O. HelinHenrik O. Helin Institute of Medical Technology, University of Tampere, Tampere , Mikael E. LundinMikael E. Lundin Biomedical Informatics Group, HUSLAB, Helsinki University Central Hospital, Helsinki , Mervi LaaksoMervi Laakso Department of Pathology, Seinäjoki Central Hospital, Seinäjoki, Finland , Johan LundinJohan Lundin Biomedical Informatics Group, HUSLAB, Helsinki University Central Hospital, Helsinki , Heikki J. HelinHeikki J. Helin Department of Oncology, University of Helsinki and Division of Pathology, HUSLAB, Helsinki University Central Hospital, Helsinki , and Jorma IsolaJorma Isola Institute of Medical Technology, University of Tampere, Tampere View All Author Informationhttps://doi.org/10.1016/S0022-5347(05)00164-3AboutAbstractPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail Abstract Purpose: Histopathological diagnosis of small focus carcinomas in prostatic needle biopsies is often assisted by IHC. To make a definitive diagnosis the pathologist must compare IHC findings with hematoxylin and eosin stained tissue morphology. We introduce what is to our knowledge a new application of virtual microscopy, in which hematoxylin and eosin, and IHC stains done sequentially on the same microscope slide can be simultaneously displayed and compared on a computer screen. Materials and Methods: A total of 30 hematoxylin and eosin stained prostatic needle biopsies were scanned with a computer controlled microscope. The slides were destained and then immunostained with a cocktail of AMACR and p63 antibodies, which labels the nuclei of nonmalignant basal cells (p63) and the cytoplasm of neoplastic glandular cells suspicious for malignancy (AMACR). The slides were then scanned again and the pairs of virtual slides were aligned for synchronized viewing. Results: The presented technique was found helpful when suspicious lesions were small and when examining the immunoprofile of specimens was warranted, in addition to examining hematoxylin and eosin stained tissue morphology. The usefulness of our approach based on virtual microscopy can be evaluated on the website http://www.webmicroscope.net/AMACRp63, which also serves as an educational tool for self-learning the correlation between hematoxylin and eosin stained tissue morphology, and AMACR/p63 IHC in prostate biopsies. Conclusions: The technology for simultaneously viewing sequentially hematoxylin and eosin and IHC stained prostate biopsies can be readily used for educational purposes, as exemplified by our website, and along with the availability of rapid virtual slide scanners it can also be used for clinical diagnostics. Histopathological diagnosis of prostatic needle biopsies can be difficult due to benign mimics of cancer, eg post-atrophic hyperplasia, mucinous metaplasia and atypical adenomatous hyperplasia, PIN and borderline changes, often termed ASAP.1–3 To better discriminate these lesions from small focus carcinomas pathologists often use IHC, in addition to standard hematoxylin and eosin staining. A central histopathological criterion is basal cells, which distinguishes benign prostate glands and PIN from cancerous glands. Basal cells can be difficult to identify with a standard morphological stain but with IHC using antibodies against high molecular weight cytokeratins4-6 and the transcription factor p637–10 their presence or absence can be ascertained. The usefulness of basal cell IHC as an aid in the interpretation of prostate biopsies is well established.4–10 Since the transcription factor p63 is a nuclear protein, it can be used simultaneously with the cytoplasmic marker AMACR (clone P504S, Dako Cytomation, Copenhagen, Denmark), which was recently discovered to be up-regulated in prostate cancer.11 An increasing number of studies have described the usefulness of AMACR IHC for the differential diagnostics of prostate carcinoma.12 More recently IHC using a cocktail of AMACR and p63 antibodies has pinpointed more specifically the nature of suspicious glands in prostatic biopsies.13,14 Positive cytoplasmic AMACR reactivity occurring without nuclear p63, ie glands lacking basal cells, gives the most comprehensive immunophenotypic evidence of carcinoma.13,14 PIN is characterized by AMACR positivity in the presence of p63 positive basal cells.13,14 Although the usefulness of AMACR and basal cell markers has been firmly documented, the concordance between IHC and a diagnosis based on hematoxylin and eosin staining is not always perfect.1,2 Therefore, the histopathological diagnosis of prostate biopsies cannot be based on IHC alone. Hematoxylin and eosin staining allows the examination of cellular morphology, especially for nucleoli, which are a central cellular criterion for carcinoma. Thus, it is well established that hematoxylin and eosin, and IHC complement each other and must be evaluated in parallel. Unfortunately suspicious lesions in prostatic needle biopsies are often small and not always present on deeper sections cut from the paraffin block for IHC.15 A solution to the problem is to routinely prepare intervening unstained slides from prostate biopsy blocks. The extra slides are stored and used for IHC should the morphological stain (hematoxylin and eosin) not provide a definitive diagnosis.16 Another approach is to destain a previously stained hematoxylin and eosin slide, followed by IHC.15,17 Due to destaining hematoxylin and eosin morphology is no longer available for comparison with IHC unless a digital imaging system is used.15 We describe an application of virtual microscopy,18–20 which allows simultaneous on-screen visualization of a hematoxylin and eosin and AMACR/p63 double IHC staining done sequentially on the same slide. The entire biopsies can be viewed at any magnification in a transparent overlay mode in a single viewing window or side by side in 2 synchronized viewing windows. This method has several practical advantages, namely in histopathology education, in second opinion and quality control situations, and in clinical diagnostics when the morphology and immunophenotype of small lesions must be compared. Materials and methods For the study we selected 23 cases with formalin fixed, paraffin embedded prostatic needle biopsies available in the archive at the Pathology Department at Central Hospital Seinäjoki, Finland. Specimens were selected that favored diagnostically challenging diagnoses, such as small focus carcinoma, PIN, ASAP, proliferative inflammatory atrophy or suspicion for carcinoma without a more detailed description. All diagnoses were based on the original hematoxylin and eosin staining only. Common diagnostic entities were also included. Ordinary 3 to 4 μm tissue sections were cut from the paraffin blocks on charged SuperFrost™ Plus slides to avoid detaching tissue sections from the slides. The slides were stained routinely with hematoxylin and eosin, and digitized using a virtual microscopy slide scanning system, as described. After slide scanning the coverslips were removed by soaking the slides in xylene until the coverslips detached. The slides were then washed with absolute ethanol, air dried and immersed in antigen retrieval buffer, composed of 0.5M tris and 1 mM ethylenediaminetetraacetic acid, pH 9. Antigen retrieval was done in an autoclave at 105C for 5 minutes, followed by a 20-minute cooling period. This procedure also removes hematoxylin and eosin staining completely. After rinsing and endogenous peroxidase quenching primary antibody incubation was done using a cocktail of antibodies to p63 (clone 4A4+Y4A3, Novocastra, Newcastle, United Kingdom), dilution 1:200, and to AMACR (clone P504S), dilution 1:200, for 30 minutes at room temperature. Antibodies were detected using a PowerVision+™ reagent kit according to manufacturer instructions. The slides were immersed for 10 minutes in diaminobenzidine and enhanced with 0.5% copper sulfate for 5 minutes. The slides were counterstained in hematoxylin, dehydrated in graded ethanols, cleared in xylene and coverslipped. The now IHC stained slides were then digitized a second time. Figure 1 shows the slide staining and scanning protocol. Fig. 1. Preparation and viewing modes of virtual slide pair. H&E stained glass slide is scanned using computer controlled microscope. Same slide is destained as result of antigen retrieval procedure and immunostained with cocktail of AMACR and p63 antibodies. IHC stained slide is scanned again, and H&E and IHC virtual slides are processed for synchronized viewing layered with blending on top of each other or side by side. Virtual microscopy An Axioskop2 MOT microscope (Zeiss GmBH, Göttingen, Germany) was equipped with a NeoFluar® oil 40× objective (numerical aperture 1.3) and motorized specimen stage. A contiguous array of digital images covering the entire biopsy was captured at 0.3 μm per pixel resolution with a color sensor camera (capture resolution 1,300 × 1,030 pixels in 3 color scanning mode). The image capture process, ie stage movement, autofocus, shading correction and image capture, was automated using KS400 software, version 3.0 (Zeiss GmBH). The acquired image files were digitally sharpened and stitched into a single montage file, which was compressed into a wavelet-type image file (enhanced compressed wavelet format) using ER Mapper software (Earth Resource Mapping Pty, West Perth, Australia). The compressed virtual slides were uploaded to a web server running Image Web Server software (Earth Resource Mapping Pty, West Perth, Australia). The virtual slide pairs (hematoxylin and eosin, and IHC) were exactly aligned for synchronized viewing by keeping the hematoxylin and eosin virtual slide as a reference and adjusting the position of the corresponding IHC virtual slide (http://www.webmicroscope.net/AMACRp63/methods). Virtual slides on the website can be viewed in a standard web browser (Microsoft® Internet Explorer or Mozilla® Firefox™) on any Windows® platform. A small (600 kb) plug-in for the browser is automatically downloaded and installed at the first viewing session. Client computer requirements are modest, that is a 1 GHz standard personal computer is recommended. Results Routinely hematoxylin and eosin stained slides were first digitized using an established virtual microscopy scanning technique. The coverslips were removed and immunohistochemical staining for AMACR and p63 was performed using a standard protocol. The antigen retrieval procedure removed the hematoxylin and eosin staining completely and the quality of immunostaining on destained slides was similar to that on ordinary unstained material. Removing the coverslip and performing AMACR/P63 immunostaining requires approximately 3 hours, followed by slide scanning, which requires 0.5 to 1 hour. Figure 1 shows the slide staining and scanning protocol. A total of 30 biopsies from 23 patients were scanned and a publicly open website containing the virtual slides was created (http://www.webmicroscope.net/AMACRp63). The virtual slides representing pairs of hematoxylin and eosin, and IHC stains can be viewed within a standard web browser (Internet Explorer or Mozilla® Firefox™). Figure 2 shows a screen shot. Fig. 2. Screen shot of 2 slide virtual microscopy interface demonstrates prostatic biopsy stained with H&E (left) and AMACR/p63 immunostaining (right). Zooming (magnification change) and navigation in sample are synchronized, ie every movement occurs in H&E and IHC viewing windows simultaneously and similarly. AMACR positive and p63 negative glands suspicious for carcinoma. Inset (upper right) shows overview image of current position and extent of large window. Circle indicates ROI. By clicking on a biopsy thumbnail image on the website a new browser window is opened and the corresponding IHC, and hematoxylin and eosin virtual slides are loaded into a single viewing window but in separate layers. The viewing interface includes basic virtual microscopy functions, such as mouse and cursor navigation, and zooming to any magnification level. A small overview window is always present in the margin, showing the current view area and allowing rapid dragging navigation. The contrast and brightness of the virtual slide can be adjusted in the browser. Initially only hematoxylin and eosin staining is visible in the viewing window but hematoxylin and eosin, and IHC image blending can be controlled using a mouse operated slider bar. Almost perfect alignment of hematoxylin and eosin, and IHC staining was achieved, allowing comparison of hematoxylin and eosin morphology and immunophenotype even at a single cell level. As an alternative to blending hematoxylin and eosin, and IHC images, the user can choose to view hematoxylin and eosin, and IHC virtual slides side by side in 2 viewing windows (fig. 2). Also, in this mode zooming (magnification change) and navigation within the sample are synchronized, ie every movement takes place in the hematoxylin and eosin, and IHC viewing windows simultaneously and similarly. The website not only demonstrates the simultaneous viewing of sequentially stained virtual slides, but also serves as an educational tool for self-learning the correlation between prostate histopathology and the interpretation of AMACR/p63 immunostaining. To improve the pedagogic value an experienced pathologist (HJH) defined and annotated ROIs in slides with foci of particular diagnostic interest. ROIs are indicated on the screen by circles. They represent carcinomatous and suspicious glands, PIN, ASAP and post-atrophic hyperplasia. ROI indicators and the annotations are initially hidden but they can be turned on by the user. By clicking on an annotation hyperlink in the right margin the corresponding ROI is automatically zoomed into the viewing window and a textual description is displayed. A general comment on each slide is also available but not the definitive diagnoses of the case, which would require inspection of all biopsies. Depending on observer experience with prostate pathology the nature of most of these lesions can be diagnosed or suspected on hematoxylin and eosin staining. However, the aid of parallel AMACR/p63 immunostaining is already apparent after a short time of use. Discussion Immunostains for high molecular weight cytokeratin, p63 and AMACR are frequently used to aid in the interpretation of prostatic biopsies containing glands suspicious for malignancy. Despite the obvious advantages of IHC it is not 100% sensitive or specific to provide a definitive diagnosis of prostatic biopsies alone.1,2 Unfortunately biopsies often contain only a few suspicious glands, which may have been cut through and, therefore, are no longer present on deeper sections cut from the paraffin block for IHC.15 For this reason obtaining immunohistochemical information on such a lesion can be difficult. Our approach is based on virtual microscopy. An entire hematoxylin and eosin stained biopsy is digitized, destained and then after immunostaining the same slide is scanned a second time. Using virtual microscopy the suspicious glands identified by hematoxylin and eosin staining, and their AMACR/p63 immunophenotype can be compared on a computer screen more easily than with any other approach. To our knowledge this is the first demonstration of the use of whole slide virtual microscopy for real-time comparison of 2 stains made sequentially on the same slide. Feedback from clinical pathologists who tested the simultaneous hematoxylin and eosin, and IHC viewing was exclusively positive. If an analogous comparison is done using stored intervening unstained sections for IHC,16 the hematoxylin and eosin, and IHC slides must be changed back and forth in the microscope, and relocating the cells or gland of interest can be difficult and time-consuming. Another approach is destaining the hematoxylin and eosin stained slide to be used for IHC.15 This procedure is similar to ours but since the hematoxylin and eosin staining is removed, a real-time comparison of tissue morphology with IHC is not possible. A modification of this protocol, termed tissue protection immunohistochemistry, was recently introduced.17 In this method all except 1 tissue section on the slide are covered with a liquid cover glass embedding medium, which protects the sections from destaining. The uncovered section is immunostained and can be compared with the adjacent hematoxylin and eosin stained sections on the same slide.17 Although the procedure has proved usable, it has the disadvantage that most immunostaining robots (autostainers) cannot perform IHC on slides covered partially with a staining protectant. Furthermore, only adjacent sections and not exactly the same section on the slide can be compared. In our virtual microscopy based system the laboratory protocols need not be altered from those used routinely for IHC of unstained slides and the entire biopsy is available for real-time comparison. Since the alignment of the hematoxylin and eosin, and IHC staining is done by adjusting the on-screen positioning of the virtual slides, the presented system for synchronized viewing should be applicable to any slide scanner with a compatible output image format. Along with the availability of rapid slide scanners hematoxylin and eosin stained slides can routinely be scanned within minutes, followed by IHC staining, which typically requires 2 to 4 hours, a second virtual slide scanning, requiring minutes, and processing of the virtual slide pair for on-screen viewing on the Internet, requiring 0.5 to 1 hour. The pathologist can then view the hematoxylin and eosin, and IHC virtual slide pair on the office computer screen via the Intranet (a local computer network). Overall with recent advances in microscope slide scanning techniques and Internet based image transfer technology virtual microscopy has emerged as an important new tool, especially for histopathology education and quality control.18 The main advantage of virtual microscopy compared to conventional printed or digital photomicrographs is the possibility of viewing entire specimens, ie any part of a whole slide at any magnification. This is particularly important for the evaluation of prostatic biopsies, when assessing the architectural growth pattern of the glands is decisive for the diagnosis. Slides are first inspected at relatively low magnification but the diagnosis requires the evaluation of cellular details at high magnification. We have previously applied virtual microscopy to assess observer variation in Gleason grading.19 An excellent agreement between Gleason grades assessed from conventional glass slides and grades assessed from virtual slides indicated that virtual slides of prostatic biopsies can be used without compromising grading accuracy.19 The web based viewing technology allows the distribution of specimens to an unlimited number of pathologists for educational and quality control purposes as well as for a second opinion in problem cases. The application presented adds simultaneous viewing of 2 slides as a new functionality that cannot be done with a conventional microscope. Conclusions The presented technique for simultaneous viewing of hematoxylin and eosin staining, and immunostaining in prostate needle biopsies can readily be applied in pathology education, and in second opinion and quality control situations. With the availability of new, more rapid slide scanners the technique could also be used in clinical diagnostics. The usefulness of the virtual microscopy based approach can be evaluated on our publicly open website (http://www.webmicroscope.net/AMACRp63). In addition to prostate biopsy pathology, the technology described can be applied in any field of histopathology in which a comparison of tissue morphology and immunophenotype at high resolution is of value. References 1 : Acinar adenocarcinoma. In: WHO Classification of Tumours. Edited by . Lyon: IARC Press2004. Google Scholar 2 : Diagnosis and reporting of limited adenocarcinoma of the prostate on needle biopsy. Mod Pathol2004; 17: 307. Google Scholar 3 : Report of the Pathology Committee: false-positive and false-negative diagnoses of prostate cancer. BJU Int2003; 92: 62. Google Scholar 4 : Use of keratin 903 as an adjunct in the diagnosis of prostate carcinoma. Am J Surg Pathol1989; 13: 389. Google Scholar 5 : Usefulness of immunoperoxidase staining with high-molecular-weight cytokeratin in the differential diagnosis of small-acinar lesions of the prostate gland. Virchows Arch A Pathol Anat Histopathol1990; 417: 191. Google Scholar 6 : Utility of immunohistochemical detection of high molecular weight cytokeratin for differential diagnosis of proliferative conditions of the prostate. Int J Urol1998; 5: 237. Google Scholar 7 : p63 is a prostate basal cell marker and is required for prostate development. Am J Pathol2000; 157: 1769. Google Scholar 8 : p63 protein expression is rare in prostate adenocarcinoma: implications for cancer diagnosis and carcinogenesis. Urology2001; 58: 619. Google Scholar 9 : Comparison of the basal cell-specific markers, 34betaE12 and p63, in the diagnosis of prostate cancer. Am J Surg Pathol2002; 26: 1161. Google Scholar 10 : Diagnostic utility of immunohistochemical staining for p63, a sensitive marker of prostatic basal cells. Mod Pathol2002; 15: 1302. Google Scholar 11 : Identification of differentially expressed genes in human prostate cancer using subtraction and microarray. Cancer Res2000; 60: 1677. Google Scholar 12 : Alpha-methylacyl CoA racemase (P504S): overview and potential uses in diagnostic pathology as applied to prostate needle biopsies. J Clin Pathol2003; 56: 892. Google Scholar 13 : Use of p63/P504S monoclonal antibody cocktail in immunohistochemical staining of prostate tissue. Appl Immunohistochem Mol Morphol2004; 12: 75. Google Scholar 14 : Diagnostic utility of a p63/alpha-methyl-CoA-racemase (p504s) cocktail in atypical foci in the prostate. Mod Pathol2004; 17: 1180. Google Scholar 15 : Efficacy of restaining prostate needle biopsies with high-molecular weight cytokeratin. Hum Pathol2000; 31: 1155. Google Scholar 16 : Use of intervening unstained slides for immunohistochemical stains for high molecular weight cytokeratin on prostate needle biopsies. Am J Surg Pathol1999; 23: 567. Google Scholar 17 : Tissue protection immunohistochemistry: a useful adjunct in the interpretation of prostate biopsy specimens and other selected cases in which immunostains are needed on minute lesions. Am J Clin Pathol2002; 117: 194. Google Scholar 18 : Virtual microscopy. J Clin Pathol2004; 57: 1250. Google Scholar 19 : Web-based virtual microscopy in teaching and standardizing Gleason grading. Hum Pathol2005; 36: 381. Google Scholar 20 : digital atlas of breast histopathology: an application of web-based virtual microscopy. J Clin Pathol2004; 57: 1288. Google Scholar © 2006 by American Urological AssociationFiguresReferencesRelatedDetails Volume 175Issue 2February 2006Page: 495-499 Advertisement Copyright & Permissions© 2006 by American Urological AssociationKeywordsprostatealpha-methylacyl-CoA racemasebiopsytelepathologyprostatic neoplasmsMetricsAuthor Information Henrik O. Helin Institute of Medical Technology, University of Tampere, Tampere More articles by this author Mikael E. Lundin Biomedical Informatics Group, HUSLAB, Helsinki University Central Hospital, Helsinki More articles by this author Mervi Laakso Department of Pathology, Seinäjoki Central Hospital, Seinäjoki, Finland More articles by this author Johan Lundin Biomedical Informatics Group, HUSLAB, Helsinki University Central Hospital, Helsinki More articles by this author Heikki J. Helin Department of Oncology, University of Helsinki and Division of Pathology, HUSLAB, Helsinki University Central Hospital, Helsinki More articles by this author Jorma Isola Institute of Medical Technology, University of Tampere, Tampere More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...
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Background: The contribution of staining techniques in brightfield microscopy has been remarkable, considering that many of these staining techniques are still widely used for diagnostic purposes more than a century after their introduction. Each working day in laboratories around the world, millions of microscope slides stained with Hematoxylin and Eosin are prepared and viewed by pathologists as part of the diagnostic process. Materials and Methods: The study included histological sections of four groups and 3 hematoxylin solutions. Each group of hematoxylin had sections of four groups. Hence, a total number of 60 sections were made. Results: The analysis of the relationship of the following variables, namely, different alum hematoxylin concluded that Harri’s hematoxylin was superior to Mayer’s and Ehrlich hematoxylin. Conclusion: Our study has shown much promise in exploring Hematoxylin and Eosin stain as a routine staining procedure. The persistence and continuing viability and growth of Hematoxylin and Eosin morphology indicates that this simple technique continues to meet most of the requirements of not only the pathologists but also clinicians, and, let us not forget, patients.
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Hematoxylin은 한국의 남부지방에서 당뇨합병증을 치료하기 위하여 사용해오던 민간약제인 Hematoxylon campechianum의 주성분이다. 본 논문에서는 hematoxylin의 혈당저하 메커니즘을 연구하기 위하여 3개 군의 흰쥐-정상군, 당뇨군, hematoxylin 처치군-에서 분리한 대퇴신경을 대상으로 2-deoxyglucose 수송능과 인지질 대사능을 조사였다. 실험결과 hematoxylin은 당뇨군에서 혈당치를 현저하게 감소시키는 것으로 나타났다. 또한 무게단위의 기준으로 당뇨군의 신경조직의 경우, 총 인지질의 양은 20% 감소하였으나 상대적으로 phosphatidylinositide의 감소는 작은 것으로 나타났다. 이 경우, hematoxylin을 처치하면 2-[3H] myo-inositol이 총 phosphoinositids로 대사되는 비율이 증가하는 것으로 밝혀졌다. 이러한 효과는 낮은 농도의 hematoxylin 처치군보다 높은 농도의 처치군에서 훨씬 큰 것으로 나타났다. 이상의 실험결과는 hematoxylin의 당뇨개선 메커니즘이 myo-inositol 대사를 증가시킴으로써 체내 당 수송과 인지질 대사를 정상화시킨다는 점을 제시하고 있다. 따라서 본 연구결과를 토대로 저자들은 hematoxylin은 향후 당뇨 치료제로서 후보 물질이 될 수 있음을 제시하고자 한다. Hematoxylin is the main component of Hematoxylon campechianum which has been utilized in the southern provinces of Korea as a folk remedy for diabetic complications. In the present study, to investigate the hypoglycemic mechanism of hematoxylin, the 2-deoxyglucose uptake and phospholipid metabolism were examined in sciatic nerves from three groups of rats : normal control, diabetic control, diabetic hematoxylin-treated group. Hematoxylin significantly reduced blood glucose levels in diabetic control rats. On a wet weight basis, the nerves from diabetic rats showed a 20% decrease in total phospholipid from that of controls and a relative decrease in phosphatidylinositide. Hematoxylin treatment increased the incorporation rate of 2-[3H] myo-inositol into total phosphoinositids in diabetic rat. The effectiveness were more potent in higher dose hematoxylin-treated rats than lower dose hematoxylin-treated rats. These results suggest that hematoxylin increases glucose transport and lipid metabolism by partially normalizing concerned with myo-inositol metabolism in diabetic rat. Therefore we propose that hematoxylin can be a promising candidate for diabetes medication.
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Helicobacter pylori and intestinal metaplasia (IM) are readily seen in hematoxylin and eosin-stained slides of gastric and/or esophageal biopsies, yet many pathology laboratories perform routine special stains on all of these biopsies. We wished to determine if special stains are necessary for every single gastric and/or esophageal biopsy. We prospectively studied 613 gastric and/or esophageal biopsies from 494 consecutive patients. The slides were stained with hematoxylin and eosin, toluidine blue (TB) for H. pylori, and Alcian blue (AB) for IM. The hematoxylin and eosin slide was classed as positive or negative for H. pylori and IM. Then it was determined if the case needed a TB or AB stain. A total of 436 cases (71.1%) were identified as H. pylori-negative and not needing a TB stain, and none was TB+. A total of 126 (20.6%) of hematoxylin and eosin slides were inconclusive for H. pylori and were regarded as needing a TB stain. Twenty of these (15.9%) were TB+. Fifty-one biopsies (8.3%) were regarded as H. pylori+ on hematoxylin and eosin; the TB stain was also positive in 49. IM was present in 113 (18.4%) hematoxylin and eosin biopsies. Hematoxylin and eosin slides were IM-negative in 498 cases (81.2%). The AB stain revealed rare goblet cells in 3 of 498 cases (0.6%). Only one of those biopsies was esophageal, and that had one goblet cell that was missed on hematoxylin and eosin. Only 2 (0.3%) were regarded as needing an AB stain. We conclude that routine special stains for all gastric and/or esophageal biopsies are not required, and hematoxylin and eosin assessment combined with selective ordering of these stains will identify virtually all cases of H. pylori gastritis and intestinal metaplasia.
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地方性甲状腺腫は臨床, 病理学的に複雑な経過を示し, 疫学的, 病理学的発生論や治療の選択に多くの難題が残されている. 著者は本症の病期検討および妥当な病期分類がこれらの検索, 解明にきわめて有用であると着目し, 甲状腺剔出を行なった地方性甲状腺腫336例を臨床, 病理学的に精査し下記の結果をえた. 1) 地方性甲状腺腫は臨床的, 病理学的経過からStage 1;過形成期, Stage 2;腫大期, Stage 3;結節形成期と分類できた. 2) 本症は病期の進行に伴い病悩期間は長くなり, 甲状腺腫は増大し種々の局所圧迫症状をみるが, 合併症がなければ全身的, 臨床生化学的所見はほぼ正常である. 3) 臨床, 病理学的に本症はStage 1からStage 2さらにStage 3に進行し, Stage 3は終末期である. 4) 病変の占居部位はStage 1では両葉性, Stage 2では両葉性と単葉性がほぼ等しく, Stage 3では単葉性が多い点からもStageの進行度を裏付けられる. 5) 336例のうち男性39例, 女性297例, 男女比1:7.6で, 発生のピークは女性では20才から30才代, 男性は30才から40才代であった. 6) 手術適応例は若年者より成人に多く, 女性は男性より著しく多い. ヨード治療の効果が若年者ほど良好で, 男性は女性よりもヨード感受性が高いためである. 7) Stage 3の9.4%に甲状腺機能亢進症 (4.03%), 腺腫 (1.34%), 甲状腺癌 (4.03%) などの共存疾患がみられた. 8) ヨード治療はStage 1では効果的であるがStage 2では無効でStage 3に進行し, 種々の合併症を起こすこともあり, Stage 2における手術が望ましい.
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Helicohacter pylorl (H. pylorl ) Is ldentlflable in hematoxyiin‐eosin (HE)‐stalned preparations, particuularly under a prolonged hematoxylln staining condition. The stalnlng intensity of H. pylori bodies was Increased when the hemetoxylln stalnlng period was doubled from 5 min, the routine period of Meyer's hematoxyiin stalnlng, to 10 min. The stalning intenstty of the background eplthellal nuclei was only mildly increased. Hematoxylin‐eosin staining employing a prolonged hematoxylin staining period gave detedablllty of H. pylori comparable to May‐Glemsa and lmmunoperoxldase staining. The modified HE method is thus Very useful and practical for identifying H. pylori in routine gastric biopsy specimens.
Gastric biopsy
Eosin
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Acquiring information on the precise distribution of a tumor is essential to evaluate intratumoral heterogeneity. Conventional hematoxylin and eosin staining, which has been used by pathologists for more than 100 years, is the gold standard of tumor diagnosis. However, it is difficult to stain entire tumor tissues with hematoxylin and eosin and then acquire the three-dimensional distribution of cells in solid tumors due to difficulties in the staining and rinsing. In this paper, we propose a modified hematoxylin and eosin staining method, in which delipidation and ultrasound waves were applied to enhance tissue permeability and accelerate dye diffusion. This improved hematoxylin and eosin staining method is termed iHE (intact tissue hematoxylin and eosin staining). We applied the iHE method to stain intact organs of mice, which were then sectioned and imaged sequentially. The results showed that the whole tissue was stained homogeneously. Combined with micro-optical sectioning tomography (MOST), the iHE method can be used for 3D volume imaging and to evaluate the intratumoral heterogeneity of the entire tumor tissue spatially. Therefore, this method may help to accurately diagnose the invasion stage of tumors and guide clinical treatments.
Stain
Eosin
Eosin Y
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