Relevant bibliographies by topics / Immunohistochemistry

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Academic literature on the topic 'Immunohistochemistry'

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Published: 4 June 2021

Last updated: 9 June 2025

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Journal articles on the topic "Immunohistochemistry"

B. Ingle, Sachin. "Immunohistochemistry." International Journal of Current Research and Review 10, no. 11 (2018): 0–1. http://dx.doi.org/10.31782/ijcrr.2018.10115.

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Taylor, Clive R. "Immunohistochemistry." Applied Immunohistochemistry & Molecular Morphology 27, no. 5 (2019): 325–26. http://dx.doi.org/10.1097/pai.0000000000000770.

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Fuchigami, Hirobumi, Kazunobu Ohnuki, Toshihiro Suzuki, and Kazumasa Takenouchi. "Immunohistochemistry." Drug Delivery System 39, no. 3 (July 25, 2024): 218–21. http://dx.doi.org/10.2745/dds.39.218.

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Mani, Haresh, and Dani S. Zander. "Immunohistochemistry." Chest 142, no. 5 (November 2012): 1324–33. http://dx.doi.org/10.1378/chest.12-0123.

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&NA;. "Immunohistochemistry." Pathology 22 (1990): 9–10. http://dx.doi.org/10.3109/00313029009060103.

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Aryal, G. "Immunohistochemistry." Journal of Pathology of Nepal 5, no. 10 (September 14, 2015): I. http://dx.doi.org/10.3126/jpn.v5i10.15660.

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Immunohistochemistry (IHC) or immunocytochemistry is a method of localizing specific antigen in tissue or cells based on antigen antibody reaction. IHC is the way of validating morphological findings. It helps in tumor diagnosis and classification, identify prognostic and predictive markers. IHC has a long history that dates back more than 70 years, when Coons1 first developed immunofluroscence technique to detect corresponding antigen in frozen tissue section. At oxford, Taylor and Burns2 developed the first successful demonstration of antigens in routinely processed formalin fixed paraffin-embedded sections. Since the early 1990s, IHC has been applied in routine formalin fixed paraffin embedded tissue. 3-4 Validation of reagents, protocols, controls and staining results are vital steps of IHC. The basic principles and protocols for fresh-frozen tissue sections are the same as those for paraffin sections, except that the antigen retrieval and dewaxing procedures are not required for frozen tissue sections. Titrations may also differ and must be separately optimized. Basic principles: Antigen-antibody recognition is based on the three-dimensional (3D) structure of protein or some other antigen, which may be compromised by formalin-induced modification of protein conformation (“masking”) but is restored in part by Antigen retrieval. Anti-A antibody binds specifically to antigen A in the tissue section. Antigen B (B) is depicted as a second antigenic determinant that is part of the anti-A molecule; anti-B antibody, made in a second species, will bind to this determinant. Thus anti-B, the so-called secondary antibody, can be used to locate the site of binding of anti-A, the primary antibody, in a tissue section. Basic IHC procedure Antigen retrieval (AR)-Enzymatic digestion (proteinase or trypsin), heat treatment (Microwave, water bath or autoclave) Blocking of non-specific background staining Incubation with primary antibody in humidity chamber Add avidin-biotin-peroxidase complex, which binds to secondary antibody Add 3, 3’ diaminobenzidine (DAB) as a chromagen (color changing reagent), with hematoxylin (Mayer's) counterstaining Theranostic Application IHC is becoming increasingly important for the evaluation of predictive markers that can help select patients who may respond to particular targeted therapies. Some of these CD117 for GI stromal tumors, Herceptin (Genentech, South San Francisco, CA) for HER2-positive breast cancers, and rituximab for CD20-positive lymphomas.

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Dabbs, David J. "Immunohistochemistry." Pathology Case Reviews 4, no. 6 (November 1999): 229. http://dx.doi.org/10.1097/00132583-199911000-00001.

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Mani, Haresh, and Dani S. Zander. "Immunohistochemistry." Chest 142, no. 5 (November 2012): 1316–23. http://dx.doi.org/10.1378/chest.11-3327.

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Mazroa, Shireen A. "Immunohistochemistry." Egyptian Journal of Histology 35, no. 2 (June 2012): 191–97. http://dx.doi.org/10.1097/01.ehx.0000414291.44156.ef.

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Wittekind, Christian, and Andrea Tannapfel. "Immunohistochemistry." Digestion 58, no. 1 (1997): 79–81. http://dx.doi.org/10.1159/000201534.

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Dissertations / Theses on the topic "Immunohistochemistry"

King, George. "Studies of amplification methods in immunohistochemistry." Thesis, University of Aberdeen, 2001. http://digitool.abdn.ac.uk/R?func=search-advanced-go&find_code1=WSN&request1=AAIU537963.

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The aim of this project was to compare and contrast the results of three related methodologies which are of particular relevance or potential within our department -. a manually performed streptAvidin Biotin Complex (sABC/ HRP) method which for many years has served as our 'standard' immunocytochemical method; an integrated semi-automated systems from DAKO based on the TechMate 500 immunostainer and the ChemMate reagent package;. a manually performed tyramide amplification method regarded as amongst the most sensitive immunocytochemical method currently available. Antigen retrieval investigating differing durations of exposure to the proteolytic enzyme trypsin, and heat mediated antigen retrieval using citrate buffer pH6.0 or DAKO high pH antigen retrieval solution were also included as parameters for investigation. To this aim a panel of eleven specific antibodies were assessed, each of which had previously displayed individual qualities worthy of investigation -. PGP9.5, CD30, oestrogen receptor, CD3 polyclonal, CD2, CD3 monoclonal, CD4, cyclin D1, kappa immunoglobulin light chain, lambda immunoglobulin light chain and IgD immunoglobulin heavy chain. The results of this work demonstrate that each of the systems investigated has its individual merits -. The manual sABC/ HRP method is the least complicated and expensive of the methods to perform. There are significant disadvantages in its used however. Being a manual method there are many steps where individual variation in working practices within a laboratory can and do affect results. The TechMate/ ChemMate system is a more sensitive system than the manual sABC/ HRP method also providing a cleaner end result and allowing higher dilutions of primary antibody to be employed. Tyramide amplification is the most sensitive system evaluated, the antibodies in this work generally demonstrated immunostaining at higher dilutions than the other methods tested although some, in particular those directed against immunoglobulin light chains, operated better under the ChemMate/ TechMate system. On occasion specific positive results were obtained which none of the other methods tested could achieve with certain antibodies, CD3 monoclonal and to a lesser extent CD2.

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Lam, Sing-chi, and 藍承志. "A HRCT and immunohistochemistry study on bronchiectasis." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2004. http://hub.hku.hk/bib/B30402499.

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Shu, Jie. "Immunohistochemistry image analysis : protein, nuclei and gland." Thesis, University of Nottingham, 2014. http://eprints.nottingham.ac.uk/27616/.

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This thesis focus on the analysis of digitized microscopic image, especially on IHC stained colour images. The corresponding contributions focused on the automatic detection of stain colour and glands, the segmentation and quantification of cell nuclei, the analysis of liver cirrhosis and the development of a semi-automatic toolbox. Colour is the most important feature in the analysis of immunostained images. We developed a statistical colour detection model for stain colour detection based on the histograms of collected colour pixels. This is acting on the approach "what you see is what you get" which outperforms the other methods on the detection of several kinds of stain colour. Verifying the presence of nuclei and quantifying positive nuclei is the foundation of cancer grading. We developed a novel seeded nuclei segmentation method which greatly improves the segmentation accuracy and reduces both over-segmentation and under-segmentation. This method has been demonstrated to be robust and accurate in both segmentation and quantification against manual labelling and counting in the evaluation process. The analysis of gland architecture, which reflects the cancer stage, has evolved into an important aspect of cancer detection. A novel morphology-based approach has been developed to segment gland structures in H-DAB stained images. This method locates the gland by focusing on its morphology and intensity characteristics, which covers variations in stain colours in different IHC images. The evaluation results have demonstrated the improvements of accuracy and efficiency. For the successive development of three methods, we put them in a semi-automatic toolbox for the aid of IHC image analysis. It can detect different kinds of stain colour and the basic components in an IHC image. The user created models and parameters can be saved and transferred to different users for the reproduction of detection results in different laboratories. To demonstrate the flexibility of our developed stained colour detection technique, the tool has been extended to the analysis of liver cirrhosis. It is a novel method based on our statistical colour detection model which greatly improves the analysis accuracy and reduces the time cost.

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Koria, Muntaha. "Identification of PHPT1 in mouse tissues by immunohistochemistry." Thesis, Uppsala University, Department of Medical Biochemistry and Microbiology, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-7745.

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Although it has been estimated that protein histidine phosphorylation account for about 6 % of the protein phosphorylation in eukaryotic cells; the knowledge of histidine phosphorylation and dephosphorylation is still limited. Lately, studies have appeared of a mammalian 14-kDa phospho- histidine phosphatase, also named protein histidine phosphatase and molecular cloning have provided some information of its physiological role. The object of the present study was to detect the protein expression of protein histidine phosphatase, PHPT1, in mouse tissue, by using immunohistochemistry. Tissue samples from a 4-week-old mouse (heart, liver, kidney, lung, muscle, and spleen), 5-month-old mouse (testis and intestinal), 8-month-old mouse (uterus) and an embryo from 14.5 days old mouse were obtained and processed for light microscopic examination. An absorption test was also made to confirm the specificity of the antibody. The results reveal that PHPT1 is mainly expressed in epithelium, heart- and skeletal muscle. These results provide new evidences for the understanding of the function of eukaryotic histidine phosphorylation and dephosphorylation.KEYWORDSPhosphohistidine, dephosphorylation, protein histidine phosphatase, phosphohistidine phosphatase, protein phosphorylation

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Sundara, Rajan Sreekumar. "Investigating male breast cancer using transcriptomics and immunohistochemistry." Thesis, University of Leeds, 2016. http://etheses.whiterose.ac.uk/15847/.

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Background: The rare nature of male breast cancer (MBC) has led to its management being guided by the extensive research conducted in the field of female breast cancer (FBC). The aim of this study was to evaluate MBC at both protein and molecular level to improve understanding of its pathology. Methodology: Immunohistochemistry analysis was performed in MBC (n=428) TMAs for 18 biomarkers (ERα, ERβ1, ERβ2, ERβ5, Total PR, AR, CK5/6, CK14, CK18, CK19, p53, Bcl-2, Her2, E-cadherin, Ki67, Survivin, Prolactin and FOXA1). The manual scoring of ERα and Ki67 was correlated with a fully automated immunohistochemistry image analysis system (ImmunoRatio™). Finally gene expression profiling (GEP) was undertaken in matched MBC (n=15) and FBC (n=10) samples. Results: There was poor 5 year overall survival (OS) in CK18 and CK19 negative patients (p= 0.05; p= 0.003), as well as poor 10 year OS in CK19 negative patients (p= 0.002). Age (p= 0.001) and nodal status (p= 0.04) was found to be independent predictors of OS at 5 years. There was significant correlations between manual and ImmunoRatio™ ERα (ρ= 0.872; p= 0.000) and Ki67 (r= 0.675; p= 0.000) scores. However due to a low measure of agreement it was not possible to validate Ki67 scoring using ImmunoRatio™. The functional enrichment analysis of GEP data using less stringent criteria (p < 0.05) identified 735 differentially expressed genes. The data analysis showed up-regulation of genes involved in ECM synthesis, degradation and re-modelling in MBC. The end product of one of the up-regulated genes (Fibronectin (FN1)) was validated in the MBC cohort with high fibronectin expression (60%) being positively associated with nodal status and showed a trend towards poor 5 year OS (p= 0.06). Conclusion: In MBC, epithelial cytokeratins, especially CK19 was found to be of prognostic significance. The extracellular matrix remodelling associated genes were found to be up-regulated in MBC. Fibronectin, end product of one of the up-regulated gene was found to have prognostic significance in MBC.

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Café, Marçal Valéria. "Pathology and immunohistochemistry of Cheetah (Acinonyx jubatus) myelopathy /." [S.l.] : [s.n.], 2006. http://www.stub.ch/index.php?p=1&i=645.

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Rozell, Björn. "Immunohistochemical studies of the thioredoxin system." Göteborg : Dept. of Histology, University of Göteborg, 1987. http://catalog.hathitrust.org/api/volumes/oclc/17242526.html.

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Kolivras, Athanassios. "Immunohistochemistry in the histopathological diagnosis of primary scalp alopecia." Doctoral thesis, Universite Libre de Bruxelles, 2016. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/238160.

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Primary scalp alopecia is classically divided into cicatricial (scarring) and non-cicatricial (non-scarring). Challenging cases are assessed with a scalp biopsy. The use of both horizontal and vertical sections (HoVert sections) has dramatically improved the accuracy of histopathological diagnosis. In this work, we have used immunostaining to address diagnostic difficulties, which persist despite all currently available tools. We performed an immunostain panel (CD3, CD4, CD8 and CD20) in order to distinguish pattern hair loss from alopecia aerate in cases which do not have the usual peribulbar lymphocytic infiltrate and showed that CD3+ T-lymphocytes within the empty fibrous follicular tracts favor a diagnosis of alopecia areata. We performed CD123 in order to distinguish lichen planopilaris from alopecia lupus erythematosus in cases with only a superficial lymphocytic infiltrate and an uninvolved interfollicular epidermis and showed that clusters of CD123+ plasmacytoid dendritic cells favor a diagnosis of lupus erythematosus. We performed cytokeratin 15 in order to assess whether the loss of the follicular bulge stem cells has diagnostic value in cicatricial alopecia and demonstrated that the loss of cytokeratin 15+ bulge stem cells is identified in lichen planopilaris, frontal fibrosing alopecia, and lupus erythematous, so cytokeratin 15 has no diagnostic value. We have attempted to integrate the new concepts and our findings into the traditional classifications of alopecia and proposed a new diagnostic algorithm. In conclusion, immunostaining combined with HoVert grossing advances the accuracy of histopathological diagnosis of primary scalp alopecia. L’alopécie primitive du cuir chevelu est habituellement classée en cicatricielle et non-cicatricielle. Dans les cas difficiles, la biopsie du cuir chevelu peut aider au diagnostic. L’utilisation de coupes, à la fois verticales et horizontales sur le même spécimen (technique HoVert), a radicalement amélioré le diagnostic histopathologique. Dans ce travail, nous avons utilisé l’immunohistochimie pour évaluer les difficultés diagnostiques qui persistent malgré tous les outils actuels. Nous avons utilisé les CD3, CD4, CD8 et CD20 pour différencier l’alopécie androgénique de la pelade dépourvue de l’infiltrat lymphocytaire péribulbaire habituel et nous avons démontré que la présence de lymphocytes CD3+ dans les travées folliculaires fibreuses est en faveur de la pelade. Nous avons utilisé le CD123 pour différencier le lichen plan pilaire du lupus érythémateux alopécie avec infiltrat lymphocytaire superficiel et sans atteinte de l’épiderme interfolliculaire et nous avons démontré que la présence d’amas de cellules dendritiques plasmacytoïdes CD123+ est en faveur du lupus érythémateux. Nous avons utilisé la cytokératine 15 pour évaluer si la perte des cellules souches du bulge a une valeur diagnostique dans l’alopécie cicatricielle et nous avons démontré que cette perte s’observait de manière identique dans le lichen plan pilaire, l’alopécie frontale fibrosante comme dans le lupus érythémateux et n’avait donc aucune valeur diagnostique. Nous avons tenté d’intégrer les nouveaux concepts et nos données dans les classifications traditionnelles des alopécies et nous avons élaboré un nouvel algorithme diagnostique. L’association des immunomarquages avec la technique HoVert ouvre de nouvelles perspectives dans le diagnostic histopathologique des alopécies primaires du cuir chevelu. Doctorat en Sciences médicales (Médecine) info:eu-repo/semantics/nonPublished

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Holtzhausen, Wendy. "Diuretic factors controlling beetle malphighian tubules fluid secretion and immunohistochemistry /." Pretoria : [s.n.], 2006. http://upetd.up.ac.za/thesis/available/etd-07182007-164904.

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Fung, Hau-yin Kevin. "Detection of SARS-CoV in lung tissues by immunohistochemistry and in-situ hybridization /." View the Table of Contents & Abstract, 2005. http://sunzi.lib.hku.hk/hkuto/record/B32020624.

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Books on the topic "Immunohistochemistry"

Kalyuzhny, Alexander E. Immunohistochemistry. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/978-3-319-30893-7.

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Simon, Renshaw, ed. Immunohistochemistry. Bloxham: Scion, 2007.

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Chu, Peiguo. Modern immunohistochemistry. Cambridge: Cambridge University Press, 2009.

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C, Cuello A., ed. Immunohistochemistry II. Chichester: Wiley, 1993.

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Del Valle, Luis, ed. Immunohistochemistry and Immunocytochemistry. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-1948-3.

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Kalyuzhny, Alexander E., ed. Signal Transduction Immunohistochemistry. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-6759-9.

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Kalyuzhny, Alexander E., ed. Signal Transduction Immunohistochemistry. Totowa, NJ: Humana Press, 2011. http://dx.doi.org/10.1007/978-1-61779-024-9.

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Kalyuzhny, Alexander E., ed. Signal Transduction Immunohistochemistry. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-2811-9.

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A, DeLellis Ronald, ed. Advances in immunohistochemistry. New York: Raven Press, 1988.

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Lin, Fan, Jeffrey W. Prichard, Haiyan Liu, and Myra L. Wilkerson, eds. Handbook of Practical Immunohistochemistry. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-030-83328-2.

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Book chapters on the topic "Immunohistochemistry"

Ramos-Vara, José A., and Luke B. Borst. "Immunohistochemistry." In Tumors in Domestic Animals, 44–87. Hoboken, NJ, USA: John Wiley & Sons, Inc., 2016. http://dx.doi.org/10.1002/9781119181200.ch3.

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Clausen, P. P., M. Møller, B. van Deurs, and O. W. Petersen. "Immunohistochemistry." In Theory and Strategy in Histochemistry, 367–86. Berlin, Heidelberg: Springer Berlin Heidelberg, 1991. http://dx.doi.org/10.1007/978-3-642-73742-8_26.

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Chiaravalli, Anna Maria. "Immunohistochemistry." In Encyclopedia of Pathology, 1–5. Cham: Springer International Publishing, 2020. http://dx.doi.org/10.1007/978-3-319-28845-1_5085-1.

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Kigawa, Junzo, Tsunehisa Kaku, Toru Sugiyama, and Steven G. Silverberg. "Immunohistochemistry." In Atlas of Clear Cell Carcinoma of the Ovary, 35–38. Tokyo: Springer Japan, 2015. http://dx.doi.org/10.1007/978-4-431-55438-7_6.

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Azumi, Norio, and Bernard Czernobilsky. "Immunohistochemistry." In Blaustein’s Pathology of the Female Genital Tract, 1131–59. New York, NY: Springer New York, 1994. http://dx.doi.org/10.1007/978-1-4757-3889-6_26.

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Taylor, Clive R., and Richard J. Cote. "Immunohistochemistry." In Encyclopedia of Cancer, 1–6. Berlin, Heidelberg: Springer Berlin Heidelberg, 2015. http://dx.doi.org/10.1007/978-3-642-27841-9_2995-2.

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Baron, Jeffrey, Jeffrey M. Voigt, Thomas T. Kawabata, and Jan A. Redick. "Immunohistochemistry." In Regulation of Hepatic Metabolism, 87–118. Boston, MA: Springer US, 1986. http://dx.doi.org/10.1007/978-1-4684-5041-5_4.

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Davis, Barbara J. "Immunohistochemistry." In Encyclopedia of Systems Biology, 1002. New York, NY: Springer New York, 2013. http://dx.doi.org/10.1007/978-1-4419-9863-7_1594.

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Carbone, Giancarlo. "Immunohistochemistry." In Antibody Usage in the Lab, 84–124. Berlin, Heidelberg: Springer Berlin Heidelberg, 1999. http://dx.doi.org/10.1007/978-3-662-03942-7_6.

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Chiaravalli, Anna Maria. "Immunohistochemistry." In Endocrine Pathology, 422–26. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-030-62345-6_5085.

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Conference papers on the topic "Immunohistochemistry"

Qiu, Fanhao, Yangyang Zhang, Xiaoguang Guo, and Zhengxia Wang. "Weakly Supervised Virtual Immunohistochemistry Staining via Schrödinger Bridge Method." In 2024 IEEE International Conference on Bioinformatics and Biomedicine (BIBM), 2369–75. IEEE, 2024. https://doi.org/10.1109/bibm62325.2024.10822509.

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Li, Jiahan, Jiuyang Dong, Shenjin Huang, Xi Li, Junjun Jiang, Xiaopeng Fan, and Yongbing Zhang. "Virtual Immunohistochemistry Staining for Histological Images Assisted by Weakly-supervised Learning." In 2024 IEEE/CVF Conference on Computer Vision and Pattern Recognition (CVPR), 11259–68. IEEE, 2024. http://dx.doi.org/10.1109/cvpr52733.2024.01070.

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Perrin, Romain, Aurélie Leborgne, Nicolas Passat, Benoît Naegel, and Cédric Wemmert. "Multi-Scale Component-Trees for Enhanced Representation in Multiplex Immunohistochemistry Imaging." In 2024 IEEE International Symposium on Biomedical Imaging (ISBI), 1–5. IEEE, 2024. http://dx.doi.org/10.1109/isbi56570.2024.10635272.

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Li, Wentao, Xingde Luo, Ziqi Yang, Xiaofang Jia, and Zhongyu Li. "A Foundation Model for Cell Segmentation from Immunohistochemistry: A Pilot Study." In 2025 IEEE 22nd International Symposium on Biomedical Imaging (ISBI), 1–5. IEEE, 2025. https://doi.org/10.1109/isbi60581.2025.10980902.

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Long, Xi, Tianyang Wang, Yanjia Kan, Yunze Wang, Silin Chen, Albert Zhou, Xianxu Hou, and Jingxin Liu. "Pseudo Training Data Generation for Unsupervised Cell Membrane Segmentation in Immunohistochemistry Images." In 2024 IEEE International Conference on Bioinformatics and Biomedicine (BIBM), 3555–60. IEEE, 2024. https://doi.org/10.1109/bibm62325.2024.10822664.

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Zabihollahy, Fatemeh, Xiaotian Yuan, Maxim Mohareb, Dexter Boehm-North, Lampros Dimitrakopoulos, Collins Wangulu, Ioannis Prassas, Neil E. Fleshner, Hong Chang, and George M. Yousef. "Automated quantification of TP53 using digital immunohistochemistry for acute myeloid leukemia prognosis." In Digital and Computational Pathology, edited by John E. Tomaszewski and Aaron D. Ward, 13. SPIE, 2025. https://doi.org/10.1117/12.3046351.

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Chen, Pingjun, Maria E. Salvatierra, Alejandra Serrano, Karina Pinao Gonzalez, Sara Ranjbar, Yasin Shokrollahi, Tanishq Gautam, et al. "Optimizing Cell Recognition in Sarcoma Multiplex Immunohistochemistry Images via Pre-Training Strategies." In 2025 IEEE 22nd International Symposium on Biomedical Imaging (ISBI), 1–5. IEEE, 2025. https://doi.org/10.1109/isbi60581.2025.10980737.

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Kawai, Masataka. "Exclusive loss in segmentation for the virtual multiplex immunohistochemistry using multiple singleplex masks." In Digital and Computational Pathology, edited by John E. Tomaszewski and Aaron D. Ward, 25. SPIE, 2025. https://doi.org/10.1117/12.3044651.

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Hang Chang, Rosa Anna DeFilippis, Thea D. Tlsty, and Bahrain Parvin. "Scoring histological sections through immunohistochemistry." In 2008 5th IEEE International Symposium on Biomedical Imaging (ISBI 2008). IEEE, 2008. http://dx.doi.org/10.1109/isbi.2008.4541003.

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Solic, Ivana, and Katarina Vukojevic. "In Search of Immunohistochemistry Quantification." In 2021 6th International Conference on Smart and Sustainable Technologies (SpliTech). IEEE, 2021. http://dx.doi.org/10.23919/splitech52315.2021.9566462.

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Reports on the topic "Immunohistochemistry"

Pleva, Christina M., Tracey A. Hamilton, John P. Petrali, and Robert K. Kan. Determining Optimal Microwave Antigen Retrieval Conditions for Microtubule-Associated Protein 2 Immunohistochemistry in the Guinea Pig Brain. Fort Belvoir, VA: Defense Technical Information Center, December 2002. http://dx.doi.org/10.21236/ada417833.

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มณีศรี เลอกรองด์, ศุภางค์. โครงการวิจัยผลของการได้รับยาพาราเซตตามอลอย่างเรื้อรังต่อการเปลี่ยนแปลงของเซลล์บุผนังหลอดเลือดสมอง : โครงสร้างในระดับจุลทรรศน์อิเลคตรอน และการหลั่งสารกระตุ้นการอักเสบ : รายงานการวิจัยฉบับสมบูรณ์. จุฬาลงกรณ์มหาวิทยาลัย, 2010. https://doi.org/10.58837/chula.res.2010.19.

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การศึกษาครั้งนี้มีจุดประสงค์เพื่อศึกษาผลของการได้รับยาพาราเซลตามอลแบบต่อเนื่องต่อการเปลี่ยนแปลงหลอดเลือดสมองและการแสดงออกของสารกระตุ้นการอักเสบ TNF alpha และIL-1 ในสมอง โดยแบ่งการทดลองออกเป็นกลุ่มควบคุม และกลุ่มที่ได้รับยาพาราเซตตามอล โดยหนูในกลุ่มที่ได้รับพาราเซตตามอลได้รับการฉีดพาราเซตตามอล 200 มิลลิกรัมต่อกิโลกรัมน้ำหนักตัวต่อเนื่องทุกวันเป็นระยะเวลา 30 วัน ในขณะที่กลุ่มควบคุมได้รับ NSS แทน ทั้งสองกลุ่มทดลองแบ่งย่อยเป็นกลุ่มที่ได้รับและไม่ได้รับการกระตุ้นให้เกิดปรากฎการณ์คอร์ติคัลสเปรดดิ้งคีเพรสชั่น โดยปรากฎการณ์คอร์ติคัลสเปรดดิ้งดีเพรสชั่นนั้นถูกเหนี่ยวนำโดยการวางผลึกโพแทสเซียมคลอไรด์ขนาด 3 มิลลิกรัม ลงบนผิวสมอง สัตว์ทดลองทุกกลุ่มจะถูกนำมาวัดการแสดงออกขอสารกระตุ้นการอักเสบ TNF alpha และ IL-1 ในสมอง โดยเทคนิค immunohistochemistry และศึกษาการเปลี่ยนแปลงของหลอดเลือดสมองโดยเทคนิค Electron microscopyผลการศึกษาวิจัยแสดงให้เห็นว่าการได้รับยาพาราเซตตามอลอย่างเรื้อรังติดต่อกันเป็นเวลานาน (30 วัน) ส่งผลเสียต่อหลอดเลือดสมอง โดยพบว่ามีการตอบสนองต่อการกระตุ้นด้วยปรากฎการณ์ CSD มากกว่ากลุ่มควบคุม ดังแสดงได้จากการตรวจพบจำนวน pinocytic vesicle, microvilli ในเซลล์บุหลอดเลือดสมองเพิ่มขึ้นรวมถึง การบวมของเซลล์แอนโตรไซด์เพิ่มขึ้นในหนูกลุ่มนี้เมื่อเทียบกับกลุ่มควบคุม ความผิดปกติของหลอดเลือดสมองที่พบนั้นมีความสอดคล้องกับการพบการแสดงออกของสารกระตุ้นการอักเสบ TNF alpha และ IL-1 ในสมองเพิ่มมากขึ้นในกลุ่มหนูที่ได้รับยาพาราเซตตามอลอย่างเรื้อรัง จากผลการศึกษาดังกล่าวแสดงให้เห็นว่าการได้รับยาพาราเซตตามอลต่อเนื่องเป็นระยะเวลานานนั้นสามารถเพิ่มการเปลี่ยนแปลงของหลอดเลือดสมองจากการกระตุ้นด้วยปรากฎการณ์ CSD มากขึ้น โดยกลไกที่ทำให้เกิดการเพิ่มขึ้นของการตอบสนองนั้นน่าจะมีความเกี่ยวข้องกับการเพิ่มการแสดงออกของสารกระตุ้นการอักเสบ TNF alpha และ IL-1 ในสมอง

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แอกทอง, สิทธิพร, та อทิตยา แก้วเสมา. การพัฒนาวิธีการรักษาการบาดเจ็บของเส้นประสาทโดยการเปลี่ยนแปลงระดับการกระตุ้นของ MAPKs : รายงานการวิจัย. จุฬาลงกรณ์มหาวิทยาลัย, 2011. https://doi.org/10.58837/chula.res.2011.21.

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การบาดเจ็บของเส้นประสาทเป็นสาเหตุที่สำคัญของภาวะทุพพลภาพเนื่องจากการตายของเซลล์ประสาทและเซลล์ค้ำจุน มีหลักฐานบ่งชี้ว่า p38 MAPK น่าจะมีบทบาทสำคัญในกระบวนการดังกล่าว การศึกษานี้มีจุดประสงค์เพื่อดูการเปลี่ยนแปลงระดับการทำงานของ p38 ในปมประสาทไขสันหลังและเส้นประสาทหลังการบาดเจ็บของเส้นประสาทและทดสอบว่า p38 เกี่ยวข้องกับกระบวนการ apoptosis ในโครงสร้างเหล่านี้หรือไม่ โดยทำให้เกิดการบาดเจ็บของเส้นประสาท sciatic แบบตัดขาด (transection) ในหนู จากนั้นเมื่อครบ 2 สัปดาห์นำเนื้อเยื่อมาศึกษาระดับ p38 และโปรตีนที่เกี่ยวข้องกับ apoptosis หนูอีกส่วนถูกแบ่งเป็นกลุ่มที่ได้รับ p38 inhibitor คือ SB203580 ขนาด 200 g/kg ต่อวันฉีดเข้าช่องท้องหรือกลุ่ม inhibitor กับกลุ่มที่ได้รับตัวทำละลาย DMSO อย่างเดียวหรือกลุ่ม control เมื่อครบ 2 สัปดาห์นำเนื้อเยื่อมาศึกษาเช่นเดียวกับหนูส่วนแรก Western blot แสดงว่าสัดส่วนของ active ต่อ total p38 (p38-P/T) ที่แสดงถึงระดับการทำงานของ p38 ในปมประสาทและเส้นประสาทไม่เปลี่ยนแปลงหลังการบาดเจ็บเช่นเดียวกับ p38 substrate คือ ATF2 นอกจากนี้ยังพบการเพิ่มระดับ total caspase-3 หลังการบาดเจ็บด้วย Immunohistochemistry แสดงว่าโปรตีนเหล่านี้ส่วนใหญ่อยู่ในเซลล์ประสาทในปมประสาทและเซลล์ค้ำจุนในเส้นประสาท เมื่อให้ยายับยั้ง p38 พบว่าทำให้ลดระดับการทำงานของ p38, ATF2 และระดับ total caspase-3 โดยเฉพาะในปมประสาท ข้อมูลเหล่านี้แสดงว่า p38 น่าจะเกี่ยวข้องกับภาวะ apoptosis ในปมประสาทไขสันหลังในภาวะเส้นประสาทบาดเจ็บ ซึ่งจำเป็นต้องยืนยันโดยการศึกษาในแง่ apoptosis โดยตรง

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ปิยะธีรธิติวรกุล, สมเกียรติ, та ศิริรัตน์ เร่งพิพัฒน์. แลกติกแอซิดแบคทีเรียที่ใช้เป็นโพรไบโอติกสำหรับปลากะพงขาว Lates calcarifer : รายงานฉบับสมบูรณ์. ศูนย์เชี่ยวชาญเฉพาะทางด้านเทคโนโลยีชีวภาพทางทะเล ภาควิชาวิทยาศาสตร์ทางทะเล คณะวิทยาศาสตร์ จุฬาลงกรณ์มหาวิทยาลัย, 2005. https://doi.org/10.58837/chula.res.2005.76.

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การแยกแลกติกแอซิดแบคทีเรียจากทางเดินอาหารของปลากะพงขาว เพื่อคัดเลือกแบคทีเรียที่มีสมบัติเป็นโพรไบโอติก ด้วยเทคนิค well agar diffusion ผลพบว่า มี 5 ไอโซเลต (กำหนดให้เป็น LAB-1 - LAB-5) ที่สามารถยับยั้งการเจริญของ Aeromonas hydrophila แบคทีเรียก่อโรคในปลาได้ ผลการวิเคราะห์ปริมาณองค์ประกอบในอาหารผสมแบบเปียก พบว่า ปริมาณสารอาหารต่างๆ ในอาหารมีเพียงพอกับความต้องการของปลากะพงขาว ยกเว้นปริมาณไขมันที่ต่ำเกินไป ความเข้มข้นของ A. hydrophila ที่ทำให้ปลาตาย 50% (LC50) หลังจากที่ปลาได้รับเชื้อแล้ว 72 ชั่วโมง มีค่าเท่ากับ 7.76 log10 เซลล์/มล. และที่ 96 และ 120 ชั่วโมง มีค่าเท่ากับ 7.47 และ 7.26 log10 เซลล์/มล. ตามลำดับ ผสมอาหารกับแต่ละไอโซเลตที่คัดแยกได้ให้มีความเข้มข้น 107 เซลล์/กรัมอาหาร นำมาเลี้ยงกับปลากะพงในตู้กระจก พบว่า มีเพียง LAB-4 เท่านั้น ที่มีการเสริมการเจริญเติบโตของปลาและต้านโรคที่เกิดจาก A. hydrophila ได้อย่างมีนัยสำคัญ (p<0.05) และเมื่อเลี้ยงปลากะพงขาวในกระชัง โดยใช้ LAB-4 ผสมในอาหารปลาโดยใช้ความเข้มข้น 105 และ 107 เซลล์/กรัมอาหาร พบว่า ทุกกลุ่มทดลองมีอัตราการรอดชีวิต การเจริญเติบโตและความสามารถในการต้านโรคที่เกิดจาก A. hydrophila ไม่แตกต่างกันอย่างมีนัยสำคัญ (p>0.05) สามารถตรวจพบ LAB-4 ในลำไส้ของปลากะพงขาวทั้ง 2 กลุ่มทดลองและไม่พบ LAB-4 ในกลุ่มควบคุม ปลากะพงขาวที่ตายหลังจากการเหนี่ยวนำให้เกิดโรคด้วย A. hydrophila แสดงวิการของโรคอย่างชัดเจนและสามารถตรวจพบ A. hydrophila ในเนื้อเยื่อของปลาที่ตายด้วยเทคนิคทาง Immunohistochemistry พิสูจน์เอกลักษณ์ของ LAB-4 ด้วยการตรวจสอบรูปร่าง ลักษณะ ปริมาณกรดแลกติกด้วยเทคนิค HPLC และตรวจสอบสมบัติบางประการทางชีวเคมี รวมทั้งการหาลำดับนิวคลีโอไทด์บริเวณ 16S rDNA เพื่อใช้เทียบเคียงกับลำดับนิวคลีโอไทด์บริเวณ 16S rDNA ของ Weissella confusa ที่มีรายงานใน Genbank พบว่า ลำดับนิวคลีโอไทด์ของ LAB-4 มีความใกล้เคียงกับ Weissella sp.

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Burke, Allen P. Detection and Clinicopathologic Correlation of Human Immunodeficiency Virus (HIV-1) Nucleic Acids and Antigens in Reticuloendothelial and Central Nervous System Tissues, by Immunohistochemistry, in situ Hybridization, and Polymerase Chain Reaction. Fort Belvoir, VA: Defense Technical Information Center, September 1992. http://dx.doi.org/10.21236/ada259307.

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Patumraj, Suthiluk, and Sheepsumon Viboolvorakul. Protective effects of exercise training against vascular and neuronal dysfunction in aging brain. Department of Physiology, Faculty of Medicine, Chulalongkorn University, 2019. https://doi.org/10.58837/chula.res.2019.11.

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During advancing age, reduction of microvessels in the brain contributes insufficiency tissue perfusion. Mounting evidence indicates that microvascular deterioration in aged brain relates to oxidative stress. Nuclear factor erythroid-related factor 2 (Nrf2) plays an important role in cellular antioxidant defense. Regular physical exercise is well known to have beneficial effect to brain health, including promoted blood flow and augmented angiogenesis, in aging individuals. However, the underlying mechanism of regular physical exercise in improvement of brain microvascular density during advancing age has not been fully elucidated. This study aimed to investigate the underlying mechanism of exercise training in improvement of microvascular density associated with PI3K/Akt/Nrf2 pathway in aged rat brain. Male Wistar rats were divided into three groups; sedentary-young (SY), sedentary-age (SA) and trained-age (TA). Exercise program included swimming exercise for eight weeks. Expression of CD31 (as indicator of microvascular density) and Nrf2 were evaluated by immunohistochemistry staining. Activity of Nrf2, protein levels of phosphatidylinositol-4,5-biphosphate 3-kinase (PI3K) and phosphorylated-protein kinase B (p-Akt) in isolated brain microvessels were assessed by immunoassay. Aging (SA) induced significant reduction of brain microvascular density and expression of Nrf2, PI3K and p-Akt proteins, as well as Nrf2 activity, comparing to those of SY group. The eight-week exercise training significantly improved brain microvascular density and upregulated Nrf2, PI3K and p-Akt proteins as well as activated Nrf2 activity, than that of the age group without exercise (SA). In conclusion, exercise training can improve brain microvascular deterioration associated with PI3K/Akt/Nrf2 pathway in aging rats.

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Tummaruk, Padet. Impact of PRRS virus infection in the replacement gilts on number and development of follicles in the ovarian tissues. Chulalongkorn University, 2015. https://doi.org/10.58837/chula.res.2015.81.

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Replacement gilts are a major source of introducing the porcine reproductive and respiratory syndrome (PRRS) virus into the swine breeding herd. The objective of the present study was to determine the prevalence of PRRS virus antigen by detection in the ovary of gilts culled due to reproductive disturbances. Ovarian tissues were collected from 100 Landrace x Yorkshire crossbred slaughtered gilts in Thailand. PRRS virus antigen in the ovarian tissues were detected by using immunohistochemistry. On average, the gilts were culled at 277 days of age at a body weight of 148 kg. Of these gilts, 75 had attained puberty and 25 had not attained puberty. PRRS virus antigens were detected in the macrophages within the ovary. The histological morphology of macrophages containing the PRRS virus in ovarian tissue was similar to that found in lung tissue. Of all the ovarian tissues (n = 100), the PRRS virus was detected in 70 (70.0%) ovaries. The detection frequency of the PRRS virus in the ovarian tissue of gilts did not differ significantly among the culling reasons (P = 0.496). The gilts that had exhibited standing oestrus had a lower PRRS virus detection than those that had never exhibited standing oestrus (62.8% versus 92.0%, respectively, P = 0.005). Gilts with a body weight of 151 – 160 kg had a higher frequency of PRRS virus detection (89.5%) than gilts with a body weight of ≤130 kg (52.9%, P = 0.025) and 131–140 kg (50.0%, P = 0.032). The ovulation rate in the ovary containing the PRRS virus tended to be lower than the ovary without the PRRS virus (14.8 and 16.7 CL, respectively, P = 0.196). These findings indicated that the natural exposure to PRRS virus in the replacement gilts caused an existence of the virus in the ovarian tissues and may subsequently influence the ovarian function.

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Suriyaphol, Gunnaporn. Study the gene expression of E-cadherin, syndecan1, matrix metalloproteinases-2, -7, -9, -14 and tissue inhibitors of metalloproteinases-1 and -2 in canine oral melanoma. Chulalongkorn University, 2015. https://doi.org/10.58837/chula.res.2015.80.

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The objectives of this study were to 1.) select the suitable reference genes for quantitative real-time polymerse chain reaction in the most common canine oral cancers: oral melanoma (OM) and oral squamous cell carcinoma (OSCC), 2.) study the gene expression of E-cadherin (CDH1), syndecan 1 (SDC1), matrix metalloproteinases-2, -7, -9, -14 (MMP2, MMP7, MMP9, MMP14) and tissue inhibitors of metalloproteinases-1 and -2 (TIMP1, TIMP2) in canine OM at the mRNA level and study the CDH1, SDC1 and Ki-67 protein expression by immunohistochemistry, and 3.) study the association of gene expression and the tumor, node, metastasis (TNM) stage. Twelve OM tissues with different TNM stages together with 7 OSCC and 8 normal gingival tissues were collected for reference gene selection. Five algorithms were used to evaluate 6 candidate reference genes, including beta actin (ACTB), beta-2-microglobulin (B2M), glyceraldehydes-3-phosphate dehydrogenase (GAPDH), ribosomal protein L13a (RPL13a), ribosomal protein S5 (RPS5), and ribosomal protein S19 (RPS19). The result showed that the cohort of the most suitable reference genes for canine OM and OSCC were ACTB, RPS5 and RPS19 and they were then used to normalize target genes in qRT-PCR. In OM, CDH1 and SDC1, encoding tumor suppressor proteins, were found to be down-regulated, corresponding to the IHC results. MMP7 gene was also found to be down-regulated whereas MMP2 and MMP14, encoding the proMMP2 activator, were upregulated, indicating cancer progression. TIMP1, encoding an inhibitor of tumor progression and metastasis, and TIMP2, encoding an inhibitor of MMP2, were upregulated in the early stage (stages 1 and 2) and late stage (stages 3-4) of the disease, respectively. Ki-67 expression, an indicator of proliferating cells, was also found to be upregulated in OM. In conclusion, the present study reported the suitable reference genes for canine OM and OSCC. The association of tumor suppressing gene and some MMP gene expression with OM was presented, regardless of the disease stages. Upregulation of TIMPs in the different stages of OM should be further studied for the possibility to be used as pathological and diagnostic markers of the disease stages.

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Funkenstein, Bruria, and Cunming Duan. GH-IGF Axis in Sparus aurata: Possible Applications to Genetic Selection. United States Department of Agriculture, November 2000. http://dx.doi.org/10.32747/2000.7580665.bard.

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Many factors affect growth rate in fish: environmental, nutritional, genetics and endogenous (physiological) factors. Endogenous control of growth is very complex and many hormone systems are involved. Nevertheless, it is well accepted that growth hormone (GH) plays a major role in stimulating somatic growth. Although it is now clear that most, if not all, components of the GH-IGF axis exist in fish, we are still far from understanding how fish grow. In our project we used as the experimental system a marine fish, the gilthead sea bream (Sparus aurata), which inhabits lagoons along the Mediterranean and Atlantic coasts of Europe, and represents one of the most important fish species used in the mariculture industry in the Mediterranean region, including Israel. Production of Sparus is rapidly growing, however, in order for this production to stay competitive, the farming of this fish species has to intensify and become more efficient. One drawback, still, in Sparus extensive culture is that it grows relatively slow. In addition, it is now clear that growth and reproduction are physiological interrelated processes that affect each other. In particular sexual maturation (puberty) is known to be closely related to growth rate in fish as it is in mammals, indicating interactions between the somatotropic and gonadotropic axes. The goal of our project was to try to identify the rate-limiting components(s) in Sparus aurata GH-IGF system which might explain its slow growth by studying the ontogeny of growth-related genes: GH, GH receptor, IGF-I, IGF-II, IGF receptor, IGF-binding proteins (IGFBPs) and Pit-1 during early stages of development of Sparus aurata larvae from slow and fast growing lines. Our project was a continuation of a previous BARD project and could be divided into five major parts: i) obtaining additional tools to those obtained in the previous project that are necessary to carry out the developmental study; ii) the developmental expression of growth-related genes and their cellular localization; iii) tissue-specific expression and effect of GH on expression of growth-related genes; iv) possible relationship between GH gene structure, growth rate and genetic selection; v) the possible role of the IGF system in gonadal development. The major findings of our research can be summarized as follows: 1) The cDNAs (complete or partial) coding for Sparus IGFBP-2, GH receptor and Pit-1 were cloned. Sequence comparison reveals that the primary structure of IGFBP-2 protein is 43-49% identical to that of zebrafish and other vertebrates. Intensive efforts resulted in cloning a fragment of 138 nucleotides, coding for 46 amino acids in the proximal end of the intracellular domain of GH receptor. This is the first fish GH receptor cDNA that had been cloned to date. The cloned fragment will enable us to complete the GH - receptor cloning. 2) IGF-I, IGF-II, IGFBP-2, and IGF receptor transcripts were detected by RT-PCR method throughout development in unfertilized eggs, embryos, and larvae suggesting that these mRNAs are products of both the maternal and the embryonic genomes. Preliminary RT-PCR analysis suggest that GH receptor transcript is present in post-hatching larvae already on day 1. 3) IGF-1R transcripts were detected in all tissues tested by RT-PCR with highest levels in gill cartilage, skin, kidney, heart, pyloric caeca, and brain. Northern blot analysis detected IGF receptor only in gonads, brain and gill cartilage but not in muscle; GH increased slightly brain and gill cartilage IGF-1R mRNA levels. 4) IGFBP-2 transcript were detected only in liver and gonads, when analyzed by Northern blots; RT-PCR analysis revealed expression in all tissues studied, with the highest levels found in liver, skin, gonad and pyloric caeca. 5) Expression of IGF-I, IGF-II, IGF-1R and IGFBP-2 was analyzed during gonadal development. High levels of IGF-I and IGFBP-2 expression were found in bisexual young gonads, which decreased during gonadal development. Regardless of maturational stage, IGF-II levels were higher than those of IGF-L 6) The GH gene was cloned and its structure was characterized. It contains minisatellites of tandem repeats in the first and third introns that result in high level of genetic polymorphism. 7) Analysis of the presence of IGF-I and two types of IGF receptor by immunohistochemistry revealed tissue- and stage-specific expression during larval development. Immunohistochemistry also showed that IGF-I and its receptors are present in both testicular and ovarian cells. Although at this stage we are not able to pinpoint which is the rate-limiting step causing the slow growth of Sparus aurata, our project (together with the previous BARD) yielded a great number of experimental tools both DNA probes and antibodies that will enable further studies on the factors regulating growth in Sparus aurata. Our expression studies and cellular localization shed new light on the tissue and developmental expression of growth-related genes in fish.

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Fields, Michael J., Mordechai Shemesh, and Anna-Riitta Fuchs. Significance of Oxytocin and Oxytocin Receptors in Bovine Pregnancy. United States Department of Agriculture, August 1994. http://dx.doi.org/10.32747/1994.7568790.bard.

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Oxytocin has multiple actions in bovine reproductive tract and it was our purpose to determine the nature of these actions and their significance for the physiology of bovine reproduction. The bovine oxytocin receptors (OTR) gene was cloned and its expression studied during the cycle and pregnancy. OTR mRNA changed in parallel with OTR with control occurring mainly at the transcriptional level. However, the endocrine regulation of OTR were found in endometrium and cervical mucosa at estrus and at parturition. In both tissues OTR were suppressed in the luteal phase and early pregnancy. Whereas cervical OTR remained suppressed throughout pregnancy, endometrial OTR began to increase soon after implantation and reached higher concentrations in midpregnancy than at estrus. OTR in caruncles did not increase until third trimester, and OTR in cervical mucosa, cotyledons and fetal membranes increased only at term. Myometrial OTR showed less variation and OTR were present throughout the cycle and pregnancy but increased significantly during mid- and late pregnancy. OTR were localized in endometrial epithelial cells and lumina epithelial cells of cervical mucosa as determined by immunohistochemistry. Endometrial OTR were functional throughout pregnancy and mediated PGF release from day 50 onwards in a receptor density related manner. OTR in cervical mucosa mediated PGE release both in vivo and in vitro, as shown in cyclic cows. The ontogeny of uterine OTR was studied from third trimester fetal stage until puberty. OTR were present in endometrium and cervical mucosa in high concentrations throughout this period; myometrial OTR began to increase somewhat later but also reached adult values by 6-mo of age. In the prepuberal heifers OT injections failed to initiate PGF2a, release. The influence of steroids on the effect of OT was examined. Ovariectomy and E2 were without effect, but P4 with or without E2 induced a massive PGF2a release in response to OT in spite of reduced OTR. Bovine cyclooxygenases (COX-1 and COX-2) were cloned and their expression studied in the endometrium of prepuberal heifers and pregnant cows. Untreated and E2 treated prepuberal heifers did not express COX-2 but P4 treated heifers did express the mRNA for COX-2, albeit weakly. During the second half of pregnancy COX-2 mRNA was strongly expressed in cotyledons and somewhat less in caruncles, whereas endometrium, myometrium and cervical mucosa showed only weak, if any, COX-2 mRNA under basal conditions. However, 2 h after OT injection significant increases in COX-2 mRNA were found in endometrial RNA. Thus OT is capable of inducing the expression of the inducible COX-2 gene, and hence the conversion of arachidonic acid to prostanoids. The results indicate that the functions of OT are numerous and probably essential for successful pregnancy and parturition.

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