Staining of E-selectin ligands on paraffin-embedded sections of tumor tissue.

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Carrascal, Mylène A; Talina, Catarina; Borralho, Paula; Gonçalo Mineiro, A; Henriques, Ana Raquel; Pen, Cláudia; Martins, Manuela; Braga, Sofia; Sackstein, Robert; Videira, Paula A

2018-05-02

The E-selectin ligands expressed by cancer cells mediate adhesion of circulating cancer cells to endothelial cells, as well as within tissue microenvironments important for tumor progression and metastasis. The identification of E-selectin ligands within cancer tissue could yield new biomarkers for patient stratification and aid in identifying novel therapeutic targets. The determinants of selectin ligands consist of sialylated tetrasaccharides, the sialyl Lewis X and A (sLe X and sLe A ), displayed on protein or lipid scaffolds. Standardized procedures for immunohistochemistry make use of the antibodies against sLe X and/or sLe A . However, antibody binding does not define E-selectin binding activity. In this study, we developed an immunohistochemical staining technique, using E-selectin-human Ig Fc chimera (E-Ig) to characterize the expression and localization of E-selectin binding sites on paraffin-embedded sections of different cancer tissue. E-Ig successfully stained cancer cells with high specificity. The E-Ig staining show high reactivity scores in colon and lung adenocarcinoma and moderate reactivity in triple negative breast cancer. Compared with reactivity of antibody against sLe X/A , the E-Ig staining presented higher specificity to cancer tissue with better defined borders and less background. The E-Ig staining technique allows the qualitative and semi-quantitative analysis of E-selectin binding activity on cancer cells. The development of accurate techniques for detection of selectin ligands may contribute to better diagnostic and better understanding of the molecular basis of tumor progression and metastasis.

Parvovirus infection: an immunohistochemical study using fetal and placental tissue.

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Li, Jing Jing; Henwood, Tony; Van Hal, Sebastian; Charlton, Amanda

2015-01-01

Parvovirus B19 infection causes 5% to 15% of cases of nonimmune hydrops fetalis. The aim of our study was to evaluate the use of immunohistochemistry in diagnosing parvovirus infection in fetal and placental tissue during routine fetal and perinatal autopsies. Histology slides of 20 cases of confirmed parvovirus infection were reviewed, and immunohistochemistry was applied to selected blocks of fetal and placental tissue. Immunohistochemistry was positive in all 20 cases, and histologic viral inclusions were seen in 19 cases. Immunohistochemical staining was closely correlated with histology and was more sensitive than histology in detecting virally infected cells, especially in autolyzed tissue. All cases also had confirmatory evidence of parvovirus infection by polymerase chain reaction of fetal liver and positive maternal serology, where it was available. We conclude that parvovirus immunohistochemistry is a reliable method for diagnosing parvovirus infection, especially in autolyzed tissue where histologic assessment may be suboptimal.

Production of tissue microarrays, immunohistochemistry staining and digitalization within the human protein atlas.

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Kampf, Caroline; Olsson, Ingmarie; Ryberg, Urban; Sjöstedt, Evelina; Pontén, Fredrik

2012-05-31

The tissue microarray (TMA) technology provides the means for high-throughput analysis of multiple tissues and cells. The technique is used within the Human Protein Atlas project for global analysis of protein expression patterns in normal human tissues, cancer and cell lines. Here we present the assembly of 1 mm cores, retrieved from microscopically selected representative tissues, into a single recipient TMA block. The number and size of cores in a TMA block can be varied from approximately forty 2 mm cores to hundreds of 0.6 mm cores. The advantage of using TMA technology is that large amount of data can rapidly be obtained using a single immunostaining protocol to avoid experimental variability. Importantly, only limited amount of scarce tissue is needed, which allows for the analysis of large patient cohorts (1 2). Approximately 250 consecutive sections (4 μm thick) can be cut from a TMA block and used for immunohistochemical staining to determine specific protein expression patterns for 250 different antibodies. In the Human Protein Atlas project, antibodies are generated towards all human proteins and used to acquire corresponding protein profiles in both normal human tissues from 144 individuals and cancer tissues from 216 different patients, representing the 20 most common forms of human cancer. Immunohistochemically stained TMA sections on glass slides are scanned to create high-resolution images from which pathologists can interpret and annotate the outcome of immunohistochemistry. Images together with corresponding pathology-based annotation data are made publically available for the research community through the Human Protein Atlas portal (www.proteinatlas.org) (Figure 1) (3 4). The Human Protein Atlas provides a map showing the distribution and relative abundance of proteins in the human body. The current version contains over 11 million images with protein expression data for 12.238 unique proteins, corresponding to more than 61% of all proteins

[Application of Immunohistochemistry and Immunofluorescence Staining in Detection of Phospholipase A2 Receptor on Paraffin Section of Renal Biopsy Tissue].

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Dong, Hong-rui; Wang, Yan-yan; Wang, Guo-qin; Sun, Li-jun; Cheng, Hong; Chen, Yi-pu

2015-10-01

To evaluate the application of immunohistochemistry and fluorescence staining method in the detection of phospholipase A2 receptor (PLA2R) on paraffin section of renal biopsy tissue,and to find an accurate and fast method for the detection of PLA2R in renal tissue. The PLA2R of 193 cases were detected by immunohistochemical staining,and the antigen was repaired by the method of high pressure cooker (HPC) hot repair plus trypsin repair. The 193 samples including 139 cases of idiopathic membranous nephropathy (IMN), 15 cases of membranous lupus nephritis, 8 cases of hepatitis B virus associated membranous nephropathy, 18 cases of IgA nephropathy, and 13 cases of minimal change diseases. To compare the dyeing effects, 22 paraffin sections of renal biopsy tissue of IMN cases with positive PLA2R were stained by using 4 different. of antigen repairing,which included HPC hot repair, HPC hot repair plus trypsin repair, water bath heat repair, and water bath heat repair plus trypsin repair. To compare the dyeing effects, 15 paraffin sections of renal biopsy tissue of IMN cases with positive PLA2R were stained by using 3 different. of antigen repairing,which included water bath heat repair plus trypsin repair, protease K digestion repair, and pepsin digestion repair. In 193 cases, the positive rate of PLA2R in IMN cases was 90.6% (126/139), and the other 54 patients without IMN were negative. Twenty-two IMN patients were positive for PLA2R by using the HPC heat repair plus trypsin repaire or the water bath heat repair plus trypsin repair;while only a few cases of 22 IMN cases were positive by using the HPC hot repair alone or water bath heat repair alone. Fifteen IMN patients were positive for PLA2R by using water bath heat repair plus trypsin repair,protease K digestion repair,and pepsin digestion repair, but the distribution of positive deposits and the background were different. PLA2R immunohistochemical staining can effectively identify IMN and secondary MN. For

Porcine intestinal mast cells. Evaluation of different fixatives for histochemical staining techniques considering tissue shrinkage

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J. Rieger

2013-07-01

Full Text Available Staining of mast cells (MCs, including porcine ones, is critically dependent upon the fixation and staining technique. In the pig, mucosal and submucosal MCs do not stain or stain only faintly after formalin fixation. Some fixation methods are particularly recommended for MC staining, for example the fixation with Carnoy or lead salts. Zinc salt fixation (ZSF has been reported to work excellently for the preservation of fixation-sensitive antigens. The aim of this study was to establish a reliable histological method for counting of MCs in the porcine intestinum. For this purpose, different tissue fixation and staining methods that also allow potential subsequent immunohistochemical investigations were evaluated in the porcine mucosa, as well as submucosa of small and large intestine. Tissues were fixed in Carnoy, lead acetate, lead nitrate, Zamboni and ZSF and stained subsequently with either polychromatic methylene blue, alcian blue or toluidine blue. For the first time our study reveals that ZSF, a heavy metal fixative, preserves metachromatic staining of porcine MCs. Zamboni fixation was not suitable for histochemical visualization of MCs in the pig intestine. All other tested fixatives were suitable. Alcian blue and toluidine blue co-stained intestinal goblet cells which made a prima facie identification of MCs difficult. The polychromatic methylene blue proved to be the optimal staining. In order to compare MC counting results of the different fixation methods, tissue shrinkage was taken into account. As even the same fixation caused shrinkage-differences between tissue from small and large intestine, different factors for each single fixation and intestinal localization had to be calculated. Tissue shrinkage varied between 19% and 57%, the highest tissue shrinkage was found after fixation with ZSF in the large intestine, the lowest one in the small intestine after lead acetate fixation. Our study emphasizes that MC counting results from

Immunohistochemical evaluation of molecular radiotherapy target expression in neuroblastoma tissue

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Gains, Jennifer E.; Gaze, Mark N. [University College London Hospitals NHS Foundation Trust, Department of Oncology, London (United Kingdom); Sebire, Neil J. [Great Ormond Street Hospital for Children NHS Foundation Trust, Department of Pathology, London (United Kingdom); Moroz, Veronica; Wheatley, Keith [University of Birmingham, Cancer Research UK Clinical Trials Unit, Birmingham (United Kingdom)

2018-03-15

Neuroblastoma may be treated with molecular radiotherapy, {sup 131}I meta-Iodobenzylguanidine and {sup 177}Lu Lutetium DOTATATE, directed at distinct molecular targets: Noradrenaline Transporter Molecule (NAT) and Somatostatin Receptor (SSTR2), respectively. This study used immunohistochemistry to evaluate target expression in archival neuroblastoma tissue, to determine whether it might facilitate clinical use of molecular radiotherapy. Tissue bank samples of formalin fixed paraffin embedded neuroblastoma tissue from patients for whom clinical outcome data were available were sectioned and stained with haematoxylin and eosin, and monoclonal antibodies directed against NAT and SSTR2. Sections were examined blinded to clinical information and scored for the percentage and intensity of tumour cells stained. These data were analysed in conjunction with clinical data. Tissue from 75 patients was examined. Target expression scores varied widely between patients: NAT median 45%, inter-quartile range 25% - 65%; and SSTR2 median 55%, interquartile range 30% - 80%; and in some cases heterogeneity of expression between different parts of a tumour was observed. A weak positive correlation was observed between the expression scores of the different targets: correlation coefficient = 0.23, p = 0.05. MYCN amplified tumours had lower SSTR2 scores: mean difference 23% confidence interval 8% - 39%, p < 0.01. Survival did not differ by scores. As expression of both targets is variable and heterogeneous, imaging assessment of both may yield more clinical information than either alone. The clinical value of immunohistochemical assessment of target expression requires prospective evaluation. Variable target expression within a patient may contribute to treatment failure. (orig.)

Effect of image compression and scaling on automated scoring of immunohistochemical stainings and segmentation of tumor epithelium

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Konsti Juho

2012-03-01

Full Text Available Abstract Background Digital whole-slide scanning of tissue specimens produces large images demanding increasing storing capacity. To reduce the need of extensive data storage systems image files can be compressed and scaled down. The aim of this article is to study the effect of different levels of image compression and scaling on automated image analysis of immunohistochemical (IHC stainings and automated tumor segmentation. Methods Two tissue microarray (TMA slides containing 800 samples of breast cancer tissue immunostained against Ki-67 protein and two TMA slides containing 144 samples of colorectal cancer immunostained against EGFR were digitized with a whole-slide scanner. The TMA images were JPEG2000 wavelet compressed with four compression ratios: lossless, and 1:12, 1:25 and 1:50 lossy compression. Each of the compressed breast cancer images was furthermore scaled down either to 1:1, 1:2, 1:4, 1:8, 1:16, 1:32, 1:64 or 1:128. Breast cancer images were analyzed using an algorithm that quantitates the extent of staining in Ki-67 immunostained images, and EGFR immunostained colorectal cancer images were analyzed with an automated tumor segmentation algorithm. The automated tools were validated by comparing the results from losslessly compressed and non-scaled images with results from conventional visual assessments. Percentage agreement and kappa statistics were calculated between results from compressed and scaled images and results from lossless and non-scaled images. Results Both of the studied image analysis methods showed good agreement between visual and automated results. In the automated IHC quantification, an agreement of over 98% and a kappa value of over 0.96 was observed between losslessly compressed and non-scaled images and combined compression ratios up to 1:50 and scaling down to 1:8. In automated tumor segmentation, an agreement of over 97% and a kappa value of over 0.93 was observed between losslessly compressed images and

Detection and Cellular Localization of Phospho-STAT2 in the Central Nervous System by Immunohistochemical Staining

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Khorooshi, Reza; Owens, Trevor

2013-01-01

Phosphorylation of signal transducers and activators of transcription (STATs) indicates their involvement in active signaling. Here we describe immunohistochemical staining procedures for detection and identification of the cellular localization of phospho-STAT2 in the central nervous system (CNS...

Immunohistochemical analysis of forensic evidence from a double homicide.

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Miller, Rodney T; Grantham, Ross; Lockett, Bruce; Temple-Camp, Cynric; Pang, James

2002-12-01

We report the use of immunohistochemical staining for analysis of forensic evidence from a double homicide. A 38-year-old woman and her 7-year-old daughter were murdered by multiple blows to the head and face with a tomahawk, resulting in multiple fragments of brain tissue scattered about the murder scene. The victims' husband and father was the main suspect, who maintained that he was out of town on business during the evening of the murders. However, a shirt taken from the suspect's car on the morning after the murders (secured by the police before the suspect visited the murder scene) was found to have two small stains. DNA analysis on the stains showed the presence of the deceased wife's DNA, and immunohistochemical stains on shirt fragments conclusively documented the presence of deep central nervous system tissue, providing the critical piece of evidence needed to arrest and prosecute the suspect. This report demonstrates that shirt or similar cloth fragments can be processed into paraffin blocks and subsequently immunostained to search for and classify types of tissue fragments that may be present on the fabric.

Immunohistochemical positive stained p53 protein in bladder transitional cell carcinoma

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Halimi Monireh

2009-04-01

Full Text Available Background: Molecular genetics and immunopathologic analysis of bladder cancer have shown some abnormalities in a number of genes and proteins that have been implicated in the development and progression of such tumors, mainly in the p53 pathway. Aims: To investigate the rate of positively stained p53 protein in patients with urothelial papillary carcinoma of the bladder (UCB by immunohistochemistry and its relationship with tumor grade, gender and age of the patients. Settings and Design: During the present cross-sectional study, 100 paraffin-embedded specimens of UCB, which were provided from biopsies of the bladder by transurethral access, were immunohistochemically stained and studied for p53 protein from May 2006 to May 2007 in our referral center pathology laboratory. Materials and Methods: First, 4 µm slices of paraffin sections were provided and then stained by the avidin-biotin peroxidase method. The rate of positively stained p53 protein (defined as positive nuclear staining in over 10% of the cells was assessed. This rate was also estimated and compared between grades, genders and age-related groups (< 70 years, ≥70 years. Statistical Analysis: The χ2 , Fisher′s exact test and Mann-Whitney U test were used for comparing. Results: The overall rate of positively stained specimens was 11% for nuclear p53 protein. This rate was significantly higher in females (10/29 vs. 1/71; P < 0.001; odds ratio [OR]: 0.23; 95% confidence interval [CI]: 4.43-306.08, patients with 70 or older than 70 years (8/42 vs. 3/58; P = 0.04; OR: 0.55; 95% CI: 1.07-17.39 and in high-grade tumors (10/58 vs. 1/42; P = 0.02; OR: 0.59; 95% CI: 0.01-0.95. Conclusions: The rate of positively stained p53 protein for UCB was lower in our population. This rate was also higher in females, patients with 70 or older than 70 years and high grade of UCB.

Immunohistochemical analysis of breast tissue microarray images using contextual classifiers

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Stephen J McKenna

2013-01-01

Full Text Available Background: Tissue microarrays (TMAs are an important tool in translational research for examining multiple cancers for molecular and protein markers. Automatic immunohistochemical (IHC scoring of breast TMA images remains a challenging problem. Methods: A two-stage approach that involves localization of regions of invasive and in-situ carcinoma followed by ordinal IHC scoring of nuclei in these regions is proposed. The localization stage classifies locations on a grid as tumor or non-tumor based on local image features. These classifications are then refined using an auto-context algorithm called spin-context. Spin-context uses a series of classifiers to integrate image feature information with spatial context information in the form of estimated class probabilities. This is achieved in a rotationally-invariant manner. The second stage estimates ordinal IHC scores in terms of the strength of staining and the proportion of nuclei stained. These estimates take the form of posterior probabilities, enabling images with uncertain scores to be referred for pathologist review. Results: The method was validated against manual pathologist scoring on two nuclear markers, progesterone receptor (PR and estrogen receptor (ER. Errors for PR data were consistently lower than those achieved with ER data. Scoring was in terms of estimated proportion of cells that were positively stained (scored on an ordinal scale of 0-6 and perceived strength of staining (scored on an ordinal scale of 0-3. Average absolute differences between predicted scores and pathologist-assigned scores were 0.74 for proportion of cells and 0.35 for strength of staining (PR. Conclusions: The use of context information via spin-context improved the precision and recall of tumor localization. The combination of the spin-context localization method with the automated scoring method resulted in reduced IHC scoring errors.

TP53 Staining in Tissue Samples of Chronic Lymphocytic Lymphoma Cases: An Immunohistochemical Survey of 51 Cases

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İbrahim Kulaç

2017-03-01

Full Text Available Objective: Chronic lymphocytic leukemia (CLL is the most common lymphoproliferative disease in adults. The aim of this study is to find out if the extent of proliferation centers or the immunohistochemical expression of p53 is related to disease prognosis. Materials and Methods: In the scope of this study, 54 biopsy specimens from 51 patients (50 of lymph nodes; the others of spleen, tonsil, orbit, and liver diagnosed with CLL at the Hacettepe University Department of Pathology in 2000-2013 were reevaluated. The clinical and demographic data of the patients were obtained from our patient database. Biopsy samples were assessed semi-quantitatively for the percentage of proliferation center/total biopsy area (PC/TBA and an immunohistochemical study was performed on representative blocks of tissues for p53 expression. Results: When the patients were divided into two categories according to Rai stage as high and low (stages 0, 1, and 2 vs. stages 3 and 4, it was seen that patients with low Rai stage had a better prognosis than those with high stages (p=0.030. However, there was no statistically significant correlation between overall survival and PC/TBA ratio or p53 expression levels. Conclusion: In our cohort, PC/TBA ratio and immunopositivity of p53 did not show correlations with overall survival.

Immunohistochemical localization of androgen receptor in rat caput epididymis during postnatal development

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Sema Timurkaan

2011-09-01

Full Text Available Objectives: The aim of this study was to investigate the developmental pattern of androgen receptor (AR in caput epididymis.Materials and methods: In this study three randomly selected rats were sacrificed at ages 21, 56, 90 and 120 days old. All male rats were anesthetized with ethyl ether before killing. Then, the caput epididymides were removed and fixed in Bouin’s fixative at +4°C for 36 hour. Afterwards the tissue samples were embedded in paraffin for routine histological methods. Later the tissues were sectioned at 5μm and mounted on poly-L-lysin-coated slides. To solve the antigen masking problem, we performed microwave stimulated antigen retrieval technique before the immunohistochemical staining. Avidin-Biotin-Peroxidase Complex (ABC method was applied for immunohistochemical staining.Results: In all age groups of rats studied, positive immunohistochemical staining for the AR appeared in nuclei of epididymal cells. The staining intensity of AR positive cells did not change depending on age. In caput epididymis, immunostainable AR was found in tubular epithelial cells (principal cells, basal cells and apical cells and peritubular smooth muscle cells. The AR staining in the epithelial cells appeared to be stronger than in the peritubular smooth muscle cells. In the epithelial cells; staining intensity was stronger in principal cells than in basal cells and apical cells.Conclusion: Staining intensity of AR positive epididymal cells irrespective of age indicated the necessity of androgens for postnatal differentiation and maintaining the structure of the epididymis. Stronger staining intensity in principal cells suggested that principal cells are more sensitive to androgen stimulation. J Clin Exp Invest 2011; 2 (3: 260-266.

Automated robust registration of grossly misregistered whole-slide images with varying stains

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Litjens, G.; Safferling, K.; Grabe, N.

2016-03-01

Cancer diagnosis and pharmaceutical research increasingly depend on the accurate quantification of cancer biomarkers. Identification of biomarkers is usually performed through immunohistochemical staining of cancer sections on glass slides. However, combination of multiple biomarkers from a wide variety of immunohistochemically stained slides is a tedious process in traditional histopathology due to the switching of glass slides and re-identification of regions of interest by pathologists. Digital pathology now allows us to apply image registration algorithms to digitized whole-slides to align the differing immunohistochemical stains automatically. However, registration algorithms need to be robust to changes in color due to differing stains and severe changes in tissue content between slides. In this work we developed a robust registration methodology to allow for fast coarse alignment of multiple immunohistochemical stains to the base hematyoxylin and eosin stained image. We applied HSD color model conversion to obtain a less stain color dependent representation of the whole-slide images. Subsequently, optical density thresholding and connected component analysis were used to identify the relevant regions for registration. Template matching using normalized mutual information was applied to provide initial translation and rotation parameters, after which a cost function-driven affine registration was performed. The algorithm was validated using 40 slides from 10 prostate cancer patients, with landmark registration error as a metric. Median landmark registration error was around 180 microns, which indicates performance is adequate for practical application. None of the registrations failed, indicating the robustness of the algorithm.

High-level expression of podoplanin in benign and malignant soft tissue tumors: immunohistochemical and quantitative real-time RT-PCR analysis.

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Xu, Yongjun; Ogose, Akira; Kawashima, Hiroyuki; Hotta, Tetsuo; Ariizumi, Takashi; Li, Guidong; Umezu, Hajime; Endo, Naoto

2011-03-01

Podoplanin is a 38 kDa mucin-type transmembrane glycoprotein that was first identified in rat glomerular epithelial cells (podocytes). It is expressed in normal lymphatic endothelium, but is absent from vascular endothelial cells. D2-40 is a commercially available mouse monoclonal antibody which binds to an epitope on human podoplanin. D2-40 immunoreactivity is therefore highly sensitive and specific for lymphatic endothelium. Recent investigations have shown widespread applications of immunohistochemical staining with D2-40 in evaluating podoplanin expression as an immunohistochemical marker for diagnosis and prognosis in various tumors. To determine whether the podoplanin (D2-40) antibody may be useful for the diagnosis of soft tissue tumors, 125 cases, including 4 kinds of benign tumors, 15 kinds of malignant tumors and 3 kinds of tumor-like lesions were immunostained using the D2-40 antibody. Total RNA was extracted from frozen tumor tissue obtained from 41 corresponding soft tissue tumor patients and 12 kinds of soft tissue tumor cell lines. Quantitative real-time PCR reactions were performed. Immunohistochemical and quantitative real-time RT-PCR analyses demonstrated the expression of the podoplanin protein and mRNA in the majority of benign and malignant soft tissue tumors and tumor-like lesions examined, with the exception of alveolar soft part sarcoma, embryonal and alveolar rhabdomyosarcoma, extraskeletal Ewing's sarcoma/peripheral primitive neuro-ectodermal tumor and lipoma, which were completely negative for podoplanin. Since it is widely and highly expressed in nearly all kinds of soft tissue tumors, especially in spindle cell sarcoma, myxoid type soft tissue tumors and soft tissue tumors of the nervous system, podoplanin is considered to have little value in the differential diagnosis of soft tissue tumors.

Activity and immunohistochemical localization of porphobilinogen deaminase in rat tissues

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Jørgensen, P E; Erlandsen, E J; Poulsen, Steen Seier

2000-01-01

the activity and the immunohistochemical localization of PBGD in the following tissues of wistar female rats: brain, heart, submandibular gland, liver, kidney, pancreas, ovary, stomach, duodenum, jejunum, ileum, colon and musculature. The PBGD activity varied considerably among the tissues. It was highest...

Variability in Immunohistochemical Detection of Programmed Death Ligand 1 (PD-L1) in Cancer Tissue Types

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Scognamiglio, Giosuè; De Chiara, Anna; Di Bonito, Maurizio; Tatangelo, Fabiana; Losito, Nunzia Simona; Anniciello, Annamaria; De Cecio, Rossella; D’Alterio, Crescenzo; Scala, Stefania; Cantile, Monica; Botti, Gerardo

2016-01-01

In normal cell physiology, programmed death 1 (PD-1) and its ligand, PD-L1, play an immunoregulatory role in T-cell activation, tolerance, and immune-mediated tissue damage. The PD-1/PD-L1 pathway also plays a critical role in immune escape of tumor cells and has been demonstrated to correlate with a poor prognosis of patients with several types of cancer. However, recent reports have revealed that the immunohistochemical (IHC) expression of the PD-L1 in tumor cells is not uniform for the use of different antibodies clones, with variable specificity, often doubtful topographical localization, and with a score not uniquely defined. The purpose of this study was to analyze the IHC expression of PD-L1 on a large series of several human tumors to correctly define its staining in different tumor tissues. PMID:27213372

Histopathological Assessment and Immunohistochemical Study of Nasopharyngeal Low Grade MALT Lymphoma

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El-Hawary, A.K.

2006-01-01

Introduction: MALT lymphoma arises in a variety of body tissues, but most often in the stomach. Though relatively rare, these MALT lymphomas may arise within several sites in the head and neck, and often present diagnostic and therapeutic challenges. Immunohistochemical analysis are helpful in confirming the diagnosis between the MALT-lymphoma and the reactive lymphoid hyperplasia. MALT-type lymphoma demonstrated characteristic negative staining for CD3, CD5 and CD43, positive staining for CD20, and monotypic staining for either kappa or lambda light chain immunoglobulin markers, whereas reactive lymphoid hyperplasia all expressed Band T cell markers. Material and Methods: 41 cases of nasopharyngeal masses were obtained from the files at pathology department, Mansoura Faculty of Medicine through the period from 2002 till 2006. 31 cases were corresponded histomorphologically to low-grade B-cell lymphoma of mucosa associated lymphoid tissue (MALT) type and 10 patients with reactive lymphoid hyperplasia of the adenoid. Hematoxylin-eosin-stained slides were reviewed to confirm the diagnosis. Immunohistochemical studies were performed on formalin-fixed, paraffin-embedded sections using the labeled streptavidin-biotin-peroxidase complex method with DAB as chromogen. The following antibodies were evaluated CD20, CD3, Kappa, lambda and cytokeratin antibodies. Results: All cases of low grade MALT lymphoma show Iymphoepitheliallesion and proliferation of centrocyte like cells. 14 cases (45.1 %) show subepithelial plasma cells. Dutcher bodies were demonstrated in 10 cases (32.2%). Monocytoid B-cells were seen in 12 cases (38.7%). Six (60%) out of the ten cases of adenoids show transmigrating lymphocyte without formation of lymphoepithelial lesion. All cases with MALT-type lymphoma expressed CD20 and not CD3 whereas 10 cases of adenoid, all expressed Band T cell markers. Immunohistochemical staining showed that 31 cases of low grade MALT lymphoma were positive for immunoglobin

Sensitivity of HER-2/neu antibodies in archival tissue samples: potential source of error in immunohistochemical studies of oncogene expression.

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Press, M F; Hung, G; Godolphin, W; Slamon, D J

1994-05-15

HER-2/neu oncogene amplification and overexpression of breast cancer tissue has been correlated with poor prognosis in women with both node-positive and node-negative disease. However, several studies have not confirmed this association. Review of these studies reveals the presence of considerable methodological variability including differences in study size, follow-up time, techniques and reagents. The majority of papers with clinical follow-up information are immunohistochemical studies using archival, paraffin-embedded breast cancers, and a variety of HER-2/neu antibodies have been used in these studies. Very little information, however, is available about the ability of the antibodies to detect overexpression following tissue processing for paraffin-embedding. Therefore, a series of antibodies, reported in the literature or commercially available, were evaluated to assess their sensitivity and specificity as immunohistochemical reagents. Paraffin-embedded samples of 187 breast cancers, previously characterized as frozen specimens for HER-2/neu amplification by Southern blot and for overexpression by Northern blot, Western blot, and immunohistochemistry, were used. Two multitumor paraffin-embedded tissue blocks were prepared from the previously analyzed breast cancers as a panel of cases to test a series of previously studied and/or commercially available anti-HER-2/neu antibodies. Immunohistochemical staining results obtained with 7 polyclonal and 21 monoclonal antibodies in sections from paraffin-embedded blocks of these breast cancers were compared. The ability of these antibodies to detect overexpression was extremely variable, providing an important explantation for the variable overexpression rate reported in the literature.

Connective tissue growth factor immunohistochemical expression is associated with gallbladder cancer progression.

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Garcia, Patricia; Leal, Pamela; Alvarez, Hector; Brebi, Priscilla; Ili, Carmen; Tapia, Oscar; Roa, Juan C

2013-02-01

Gallbladder cancer (GBC) is an aggressive neoplasia associated with late diagnosis, unsatisfactory treatment, and poor prognosis. Molecular mechanisms involved in GBC pathogenesis remain poorly understood. Connective tissue growth factor (CTGF) is thought to play a role in the pathologic processes and is overexpressed in several human cancers, including GBC. No information is available about CTGF expression in early stages of gallbladder carcinogenesis. Objective.- To evaluate the expression level of CTGF in benign and malignant lesions of gallbladder and its correlation with clinicopathologic features and GBC prognosis. Connective tissue growth factor protein was examined by immunohistochemistry on tissue microarrays containing tissue samples of chronic cholecystitis (n = 51), dysplasia (n = 15), and GBC (n = 169). The samples were scored according to intensity of staining as low/absent and high CTGF expressers. Statistical analysis was performed using the χ(2) test or Fisher exact probability test with a significance level of P Connective tissue growth factor expression showed a progressive increase from chronic cholecystitis to dysplasia and then to early and advanced carcinoma. Immunohistochemical expression (score ≥2) was significantly higher in advanced tumors, in comparison with chronic cholecystitis (P < .001) and dysplasia (P = .03). High levels of CTGF expression correlated with better survival (P = .04). Our results suggest a role for CTGF in GBC progression and a positive association with better prognosis. In addition, they underscore the importance of considering the involvement of inflammation on GBC development.

A new technique for Gram staining paraffin-embedded tissue.

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Engbaek, K; Johansen, K S; Jensen, M E

1979-01-01

Five techniques for Gram staining bacteria in paraffin sections were compared on serial sections of pulmonary tissues from eight bacteriological necropsies. Brown and Hopp's method was the most satisfactory for distinguishing Gram-positive and Gram-negative bacteria. However, this method cannot be recommended as the preparations were frequently overstained, and the Gram-negative bacteria were stained indistinctly. A modification of Brown and Hopps' method was developed which stains larger numbers of Gram-negative bacteria and differentiates well between different cell types and connective tissue, and there is no risk of overstaining. PMID:86548

Tissue clearing for confocal imaging of native and bio-artificial skeletal muscle.

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Decroix, L; Van Muylder, V; Desender, L; Sampaolesi, M; Thorrez, L

2015-01-01

Novel clearing techniques have revolutionized three-dimensional confocal imaging of the brain without the need for physical tissue sectioning. We evaluated three clearing methods, ScaleA2, Clear(T2), and 3DISCO for visualizing native and tissue engineered muscle by confocal microscopy. We found that Clear(T2) treatment improved the depth of visualization of immunohistochemical staining slightly, but did not improve depth of visualization of endogenous green fluorescent protein (GFP). ScaleA2 preserved endogenous GFP signal better and permitted significantly deeper GFP imaging, but it was incompatible with tropomyosin immunohistochemical staining. 3DISCO treatment preserved both endogenous GFP and immunohistochemical staining, and permitted significantly deeper imaging. Clearing time for the 3DISCO procedure is short compared to ScaleA2 and Clear(T2). We suggest that 3DISCO is the preferable clearing method for native and tissue engineered skeletal muscle tissue.

Immunohistochemical Similarities between Lichen Sclerosus et Atrophicus and Morphea: A Case Study

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Aya Kakizaki

2015-03-01

Full Text Available Both morphea and lichen sclerosus et atrophicus (LSA are connective tissue diseases that mainly affect the skin. A recent report suggested that a substantial portion of morphea coexists with LSA. In this report, we describe a case of LSA on the abdomen accompanied by morphea; we employed immunohistochemical staining for periostin as well as MMP-7 and MMP-28, both of which are reported to facilitate fibrosis in the development of various organs, including skin. To our knowledge, this is first English language paper that demonstrates the immunohistochemical staining of periostin, MMP-7 and MMP-28 for morphea and LSA. Our present case might suggest possible mechanisms for the coexistence of two different sclerotic skin disorders.

Immunohistochemical Study of Scrub Typhus: A Report of Two Cases

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Bo-Yuan Tseng

2008-02-01

Full Text Available Scrub typhus is a zoonotic disease caused by Orientia tsutsugamushi, which is transmitted by chiggers. The target cells of this rickettsia are poorly defined in humans. Immunohistochemical staining of tissue sections of patients with scrub typhus is helpful in investigating the target cells of this rickettsia in different organs. We studied two autopsy specimens by immunohistochemical staining using a specific antibody against O. tsutsugamushi. Rickettsiae were located in endothelial cells in all of the organs evaluated, namely heart, lung, brain, kidney, appendix and skin, within cardiac muscle cells and renal tubular epithelial cells, and in macrophages located in the lymph node, liver and spleen. In conclusion, O. tsutsugamushi may disseminate into multiple organs through endothelial cells and macrophages, resulting in the development of fatal complications.

Immunohistochemical profiles of claudin-3 in primary and metastatic prostatic adenocarcinoma

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Becich Michael J

2011-01-01

Full Text Available Abstract Background Claudins are integral membrane proteins that are involved in forming cellular tight junctions. One member of the claudin family, claudin-3, has been shown to be overexpressed in breast, ovarian, and pancreatic cancer. Here we use immunohistochemistry to evaluate its expression in benign prostatic hyperplasia (BPH, prostatic intraepithelial neoplasia (PIN, normal tissue adjacent to prostatic adenocarcinoma (NAC, primary prostatic adenocarcinoma (PCa, and metastatic prostatic adenocarcinoma (Mets. Methods Tissue microarrays were immunohistochemically stained for claudin-3, with the staining intensities subsequently quantified and statistically analyzed using a one-way ANOVA with subsequent Tukey tests for multiple comparisons or a nonparametric equivalent. Fifty-three cases of NAC, 17 cases of BPH, 35 cases of PIN, 107 cases of PCa, and 55 cases of Mets were analyzed in the microarrays. Results PCa and Mets had the highest absolute staining for claudin-3. Both had significantly higher staining than BPH (p Conclusions To our knowledge, this represents one of the first studies comparing the immunohistochemical profiles of claudin-3 in PCa and NAC to specimens of PIN, BPH, and Mets. These findings provide further evidence that claudin-3 may serve as an important biomarker for prostate cancer, both primary and metastatic, but does not provide evidence that claudin-3 can be used to predict risk of metastasis.

Determination of ABO blood grouping from human oral squamous epithelium by the highly sensitive immunohistochemical staining method EnVision+.

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Noda, Hiroshi; Yokota, Makoto; Tatsumi, Shinji; Sugiyama, Shizuyuki

2002-03-01

Using the highly sensitive immunohistochemical staining method EnVision+, which employs a dextran polymer reagent for the secondary antibody, the detection of the ABH antigens was attempted in the oral squamous epithelium. This new technique uses monoclonal antibody as a primary antibody and it takes about three hours for staining. The time is much shorter than conventional absorption-elution testing or absorption-inhibition testing for the determination of ABO blood grouping. Secretor saliva samples were stained at strong intensity by the antibody, which corresponded to its blood group and anti-H. On the one hand, nonsecretor saliva samples were stained at strong intensity only by the antibody that corresponded to its blood group, and at weak intensity only by anti-H. Since human oral squamous epithelium antigens were stained specifically by this method, we can examine the ABO blood group of saliva samples and perform cytodiagnosis at the same time. Our research suggested that the EnVision+ Method is a useful technique for ABO blood grouping of saliva in forensic cases.

Hirschsprung's disease diagnosis: Comparison of immunohistochemical, hematoxilin and eosin staining

OpenAIRE

Memarzadeh, Mehrdad; Talebi, Ardeshir; Edalaty, Masod; Hosseinpour, Mehrdad; Vahidi, Nasrin

2009-01-01

Background: The diagnosis of Hirschsprung's disease (HD) is based on the absence of ganglion cells. In hemotoxilin and eosin (H and E) as well as acetylcholine esterase staining there are limitations in the diagnosis of immature ganglion cells in neonates. Methods: In this prospective study, 54 biopsies taken from suspected HD patients (five mucosal specimens and 49 full thickness specimens) were studied. In the laboratory, after preparing sections of paraffin embedded tissues, H and E staini...

A novel, modernized Golgi-Cox stain optimized for CLARITY cleared tissue.

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Kassem, Mustafa S; Fok, Sandra Y Y; Smith, Kristie L; Kuligowski, Michael; Balleine, Bernard W

2018-01-15

High resolution neuronal information is extraordinarily useful in understanding the brain's functionality. The development of the Golgi-Cox stain allowed observation of the neuron in its entirety with unrivalled detail. Tissue clearing techniques, e.g., CLARITY and CUBIC, provide the potential to observe entire neuronal circuits intact within tissue and without previous restrictions with regard to section thickness. Here we describe an improved Golgi-Cox stain method, optimised for use with CLARITY and CUBIC that can be used in both fresh and fixed tissue. Using this method, we were able to observe neurons in their entirety within a fraction of the time traditionally taken to clear tissue (48h). We were also able to show for the first-time that Golgi stained tissue is fluorescent when visualized using a multi-photon microscope, allowing us to image synaptic spines with a detail previously unachievable. These novel methods provide cheap and easy to use techniques to investigate the morphology of cellular processes in the brain at a new-found depth, speed, utility and detail, without previous restrictions of time, tissue type and section thickness. This is the first application of a Golgi-Cox stain to cleared brain tissue, it is investigated and discussed in detail, describing different methodologies that may be used, a comparison between the different clearing techniques and lastly the novel interaction of these techniques with this ultra-rapid stain. Copyright © 2017 Elsevier B.V. All rights reserved.

Digital simulation of staining in histopathology multispectral images: enhancement and linear transformation of spectral transmittance.

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Bautista, Pinky A; Yagi, Yukako

2012-05-01

Hematoxylin and eosin (H&E) stain is currently the most popular for routine histopathology staining. Special and/or immuno-histochemical (IHC) staining is often requested to further corroborate the initial diagnosis on H&E stained tissue sections. Digital simulation of staining (or digital staining) can be a very valuable tool to produce the desired stained images from the H&E stained tissue sections instantaneously. We present an approach to digital staining of histopathology multispectral images by combining the effects of spectral enhancement and spectral transformation. Spectral enhancement is accomplished by shifting the N-band original spectrum of the multispectral pixel with the weighted difference between the pixel's original and estimated spectrum; the spectrum is estimated using M transformed to the spectral configuration associated to its reaction to a specific stain by utilizing an N × N transformation matrix, which is derived through application of least mean squares method to the enhanced and target spectral transmittance samples of the different tissue components found in the image. Results of our experiments on the digital conversion of an H&E stained multispectral image to its Masson's trichrome stained equivalent show the viability of the method.

AutoIHC-scoring: a machine learning framework for automated Allred scoring of molecular expression in ER- and PR-stained breast cancer tissue.

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Tewary, S; Arun, I; Ahmed, R; Chatterjee, S; Chakraborty, C

2017-11-01

In prognostic evaluation of breast cancer Immunohistochemical (IHC) markers namely, oestrogen receptor (ER) and progesterone receptor (PR) are widely used. The expert pathologist investigates qualitatively the stained tissue slide under microscope to provide the Allred score; which is clinically used for therapeutic decision making. Such qualitative judgment is time-consuming, tedious and more often suffers from interobserver variability. As a result, it leads to imprecise IHC score for ER and PR. To overcome this, there is an urgent need of developing a reliable and efficient IHC quantifier for high throughput decision making. In view of this, our study aims at developing an automated IHC profiler for quantitative assessment of ER and PR molecular expression from stained tissue images. We propose here to use CMYK colour space for positively and negatively stained cell extraction for proportion score. Also colour features are used for quantitative assessment of intensity scoring among the positively stained cells. Five different machine learning models namely artificial neural network, Naïve Bayes, K-nearest neighbours, decision tree and random forest are considered for learning the colour features using average red, green and blue pixel values of positively stained cell patches. Fifty cases of ER- and PR-stained tissues have been evaluated for validation with the expert pathologist's score. All five models perform adequately where random forest shows the best correlation with the expert's score (Pearson's correlation coefficient = 0.9192). In the proposed approach the average variation of diaminobenzidine (DAB) to nuclear area from the expert's score is found to be 7.58%, as compared to 27.83% for state-of-the-art ImmunoRatio software. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

Langerhans cells in lichen planus and lichenoid mucositis an immunohistochemical study

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M Devi

2014-01-01

Full Text Available Aim: The aim of this study is to identify and evaluate Langerhans cell (LC in lichen planus (LP, lichenoid mucositis (LM and normal mucosa (NM using CD1a monoclonal antibody immunohistochemically. Materials and Methods: A total of 15 cases of oral lichen planus and 15 cases of LM were selected based on clinical examination and confirmed by histopathological analysis. The biopsies from the 10 patients were taken from normal buccal mucosa as control. Paraffin blocks of tissue were made, which are used for routine hematoxylin and eosin staining and immunohistochemical staining using biotin streptavidin methods (CD1a monoclonal antibody. Analysis of CD1a expression was performed by evaluating the labeling index (LI for each slide. Results: The mean CD1a LI for LP was significantly higher than that of LM and NM in the basal and supra basal layer. The mean CD1a positive cells in the connective tissues for LP were higher than that of LM and NM. Conclusion: This study clearly demonstrates a statistically significant increase in number of LC in LP than in LM, indicating the possible different immunopathogenic mechanisms.

Immunohistochemical Study of Expression of Sohlh1 and Sohlh2 in Normal Adult Human Tissues.

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Xiaoli Zhang

Full Text Available The expression pattern of Sohlh1 (spermatogenesis and oogenesis specific basic helix-loop-helix 1 and Sohlh2 in mice has been reported in previous studies. Sohlh1 and Sohlh2 are specifically expressed in spermatogonia, prespermatogonia in male mice and oocytes of primordial and primary follicles in female mice. In this report, we studied the expression pattern of Sohlh1 and Sohlh2 in human adult tissues. Immunohistochemical staining of Sohlh1 and Sohlh2 was performed in 5 samples of normal ovaries and testes, respectively. The results revealed that Sohlh genes are not only expressed in oocytes and spermatogonia, but also in granular cells, theca cells, Sertoli cells and Leydig cells, and in smooth muscles of blood vessel walls. To further investigate the expression of Sohlh genes in other adult human tissues, we collected representative normal adult tissues developed from three embryonic germ layers. Compared with the expression in mice, Sohlhs exhibited a much more extensive expression pattern in human tissues. Sohlhs were detected in testis, ovary and epithelia developed from embryonic endoderm, ectoderm and tissues developed from embryonic mesoderm. Sohlh signals were found in spermatogonia, Sertoli cells and also Leydig cells in testis, while in ovary, the expression was mainly in oocytes of primordial and primary follicles, granular cells and theca cells of secondary follicles. Compared with Sohlh2, the expression of Sohlh1 was stronger and more extensive. Our study explored the expression of Sohlh genes in human tissues and might provide insights for functional studies of Sohlh genes.

Immunohistochemical Expression of Tissue Inhibitor of Metalloproteinase-1 (Timp-1 in Invasive Breast Carcinoma

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Suada Kuskunović

2009-05-01

Full Text Available Tissue inhibitor of metalloproteinase-1 (TIMP-1 is a natural inhibitor of matrix metalloproteinas-es (MMPs. Aim of this study was to assess the immunohistochemical expression of TIMP-1 in invasive breast carcinomas, and to examine its association with classical clinico-pathological parameters, oestrogen receptor, progesterone receptor and Her-2/neu protein expression. Immuno-histochemistry was used to determine the expression of TIMP-1 on 38 paraffin-embedded breast tissue specimens - 18 with invasive ductal carcinoma, 10 with invasive lobular carcinoma, and 10 specimens from patients with fibrocystic breast disease. TIMP-1 protein was immunodetected in the carcinoma cells, fibroblasts and inflammatory cells of the stroma in 92,9%, 65,8%, and 65,8% of cases, respectively. TIMP-1 protein expression in carcinoma cells showed positive correlation with TIMP-1 protein expression in peritumoural fibroblasts (p=0,010. Positive peritumoural fibroblast TIMP-1 expression was associated with histological tumour type with higher frequency in ductal carcinomas (p=0,023. Negative association was found between TIMP-1 protein expression in carcinoma cells and HER-2/neu nuclear staining (p=0,005. TIMP-1 may be particularly useful as a predictive marker in breast carcinoma when evaluated along with HER-2/neu protein being a promising indicator of favourable prognosis in breast carcinoma.

Darkfield illumination improves microscopic detection of metals in Timm's stained tissue

DEFF Research Database (Denmark)

Baatrup, E; Frederickson, C J

1989-01-01

Deposits of trace or toxic metals can be quickly identified by light microscopical surveys of tissue sections stained for metals by variants of Timm's silver enhancement method. The present work shows that the small, isolated silver grains that label isolated deposits of metal in tissue are undet...... are undetectable in brightfield light microscopy but are easily detected in darkfield microscopy. Darkfield illumination is therefore recommended for improving the detection of trace or toxic metals in tissue. Udgivelsesdato: 1989-Aug......Deposits of trace or toxic metals can be quickly identified by light microscopical surveys of tissue sections stained for metals by variants of Timm's silver enhancement method. The present work shows that the small, isolated silver grains that label isolated deposits of metal in tissue...

Histopathological and Immunohistochemical Evaluation of Pannus Tissue in Patients with Prosthetic Valve Dysfunction.

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Karakoyun, Süleyman; Ozan Gürsoy, Mustafa; Yesin, Mahmut; Kalçık, Macit; Astarcıoğlu, Mehmet Ali; Gündüz, Sabahattin; Emrah Oğuz, Ali; Çoban Kökten, Şermin; Nimet Karadayı, Ayşe; Tuncer, Altuğ; Köksal, Cengiz; Gökdeniz, Tayyar; Özkan, Mehmet

2016-01-01

Prosthetic valve dysfunction due to pannus formation is a rare but serious complication. Currently, limited data are available concerning the pathogenesis and immunohistochemical properties of pannus. The study aim was to investigate the morphological, histopathological and immunohistochemical characteristics of pannus formation in patients with prosthetic valve dysfunction. A total of 35 patients (10 males, 25 females; mean age 44 ± 16 years) who had undergone re-do valve surgery due to prosthetic valve obstruction was enrolled in the study. Immunohistochemical studies were aimed at evaluating the expression of alphasmooth muscle actin (α-SMA) and desmin in myofibroblasts and smooth muscle cells; epithelial membrane antigen (EMA) in epithelial cells; and CD34, Factor VIII and vascular endothelial growth factor (VEGF) in endothelial cells. Matrix metalloproteinases (MMPs) -2 and -9, and transforming growth factor-beta (TGF-β) were used to demonstrate cytokine release from macrophages, leukocytes, fibroblasts and myofibroblasts. Pannus appeared as a tough and thick tissue hyperplasia which began from outside the suture ring in the periannular region and extended to the inflow and outflow surfaces of the prosthetic valves. Histopathological analysis showed the pannus tissue to consist of chronic inflammatory cells (lymphocytes, plasma cells, macrophages and foreign body giant cells), spindle cells such as myofibroblasts, capillary blood vessels and endothelial cells laying down the lumens. Calcification was present in the pannus tissue of 19 explanted prostheses. Immunohistochemical studies revealed positive α-SMA expression in all patients, whereas 60.5% of patients were positive for desmin, 50% for EMA, 42.1% for VEGF, 39.5% for TBF-β, 42.1% for MMP-2, 86.8% for CD34, and 97.4% for Factor VIII. MMP-9 was negative in all patients. Pannus tissue appears to be formed as the result of a neointimal response in periannular regions of prosthetic valves that consist

Collagenase-3 expression in periapical lesions: an immunohistochemical study.

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Bhalla, G; Astekar, M S; Ramesh, G; Kaur, P; Sowmya, G V

2014-08-01

Collagenase-3 (matrix metalloproteinase-13) is a metalloproteinase (MMP) that is associated with bone lesions and exhibits variable expression patterns in odontogenic cysts; it may play a role in regulating focal proliferation and maturation of jaw cyst epithelium. We studied the localization, staining intensity and distribution of collagenase-3 in 13 periapical granulomas with epithelium, 16 periapical granulomas without epithelium and 10 radicular cysts using archived formalin fixed, paraffin embedded tissues. A monoclonal antibody against human collagenase-3 was used to evaluate its expression. Immunohistochemical staining intensities of collagenase-3 in all periapical lesions were (-), 4 (10%); (+), 1 (3%); (++), 22 (56%) and (+++), 12 (31%); differences were not statistically significant. Immunohistochemical distribution of collagenase-3 in epithelial cells was (-), 17 (44%); (+), 17 (44%); (++), 5 (13%); in fibroblasts it was (-), 8 (20%); (+), 23 (59%); (++), 8 (21%); in plasma cells it was (-), 7 (18%); (+), 22 (56%); (++), 10 (26%); in macrophages it was (-), 7 (18%); (+), and 15 (38%); and (++), 17 (44%). Statistically significant differences were found in epithelial cells (p = 0.00) and fibroblasts (p = 0.02), whereas differences were not statistically significant for plasma cells and macrophages. Collagenase-3 may play a role in the conversion of a periapical granuloma with epithelium to radicular cyst. MMP's influence not only epithelial rest cell migration, but also invasion of various stromal cells into granulomatous tissue.

[Histopathological and immunohistochemical studies on mucous cysts].

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Kuroda, N

1989-01-01

The present study investigated the histopathology, histochemistry of mucopolysaccharides, and immunohistochemistry of oral mucous cysts. The materials were obtained from ninety cases that were histopathologically diagnosed as oral mucous cysts at the Department of Oral Pathology, Meikai University School of Dentistry. Mucopolysaccharide staining was done with PAS, alcian blue (AB, pH 2.5) and high iron diamine (HID). Immunohistochemical studies were focused on secretory component (SC), lactoferrin (Lf), alpha-amylase (Am), IgA, lysozyme (Ly), and keratin (Kr). The following results were obtained: 1. Histopathological findings. (1) Retention and/or retention-like type cysts occurred in was twenty-six cases and the extravasation type in sixty-four cases. (2) Cases showing epithelial lining of the cystic wall were only eight in number, and many cystic walls were contained granulation tissue (fifty cases). (3) As for inflammation of the cystic wall, the degree was slight, and infiltrated cells were mainly macrophages (so-called mucinophages) and lymphocytes. (4) Regarding adjoining salivary glands, acinar cells showed atrophic changes, and hypertrophy of mucous acinar cells was evident. Many ducts showed dilatation, and stromal connective tissue showed fibrosis and hyalinization. 2. Histochemical findings on mucopolysaccharides. (1) Mucous materials in cystic cavity, mucous acinar cells, and secretory materials in ductal lumens were intensely stained by PAS and AB. But stainability with AB was less than that with PAS staining. Serous acinar cells and ductal epithelium were negative to PAS and AB staining. (2) Stainability of the above with HID was less than at with PAS or AB. Cystic walls were not stained by HID. Mucous acinar cells reactive with HID were intensely stained, but the number of the positive cells was limited when compared with the numbers of PAS-and AB-positive cells. 3. Immunohistochemical findings. (1) As for mucous materials in the cystic cavity

Immunohistochemical/histochemical double staining method in the study of the columnar metaplasia of the oesophagus

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D. Cabibi

2014-03-01

Full Text Available Intestinal metaplasia in Barrett’s oesophagus (BO represents an important risk factor for oesophageal adenocarcinoma. Instead, few and controversial data are reported about the progression risk of columnar-lined oesophagus without intestinal metaplasia (CLO, posing an issue about its clinical management. The aim was to evaluate if some immunophenotypic changes were present in CLO independently of the presence of the goblet cells. We studied a series of oesophageal biopsies from patients with endoscopic finding of columnar metaplasia, by performing some immunohistochemical stainings (CK7, p53, AuroraA combined with histochemistry (Alcian-blue and Alcian/PAS, with the aim of simultaneously assess the histochemical features in cells that shows an aberrant expression of such antigens. We evidenced a cytoplasmic expression of CK7 and a nuclear expression of Aurora A and p53,  both in goblet cells of BO and in non-goblet cells of CLO, some of which showing mild dysplasia. These findings suggest that some immunophenotypic changes are present in CLO and they can precede the appearance of the goblet cells or can be present independently of them, confirming the conception of BO as the condition characterized by any extention of columnar epithelium. This is the first study in which a combined immunohistochemical/histochemical method has been applied to Barrett pathology.

Immunohistochemical comparison of markers for wound healing on plastic-embedded and frozen mucosal tissue.

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Mai, Ronald; Gedrange, Tomasz; Leonhardt, Henry; Sievers, Nicole; Lauer, Günter

2009-01-01

Immunohistologic investigations of wound healing in human oral mucosa require specific cell biological markers as well as consecutive small biopsies. Small specimens are ideally embedded in plastic (methylmethacrylate, MMA) resin due to their miniature size. This limits the use of antibodies for these markers. In this immunohistochemical study, the distribution of wound healing markers, e.g. cytokeratin (CK), laminin, collagen IV, vimentin, vinculin and fibronectin, were compared between semithin sections of plastic-embedded tissue and frozen sections of mucosal tissue in order to assess their use for future investigations. The antibodies against laminin, collagen IV and CK 1/2/10/11, 5/6, 13, 14, 17, 19 gave comparable staining patterns on cryostat sections of attached mucosa and on semithin sections of MMA-embedded attached mucosa. In the epithelial cell layers, the following distribution of CK immunostaining was observed: The basal cell layer was positive for CK 5/6, CK 14 and CK 19; the intermediate cell layer for CK 13, CK 17 and CK 1/2/10/11, and the superficial cell layer for CK 13 and CK 1/2/10/11. For most of these antibodies, enzyme digestion with 0.1% trypsin was adequate for demasking the antigens, except for anti-CK 14, anti-CK 17 and anti-laminin; predigestion with 0.4% pepsin in 0.01 N HCl gave similar staining results. The antibodies against vimentin, vinculin, fibronectin and CK 4 showed no affinity or a reciprocal reaction on the semithin sections. Therefore, the antibodies against CK 1/2/10/11; 5/6; 13; 14; 17, and 19, as well as the basement proteins laminin and collagen IV are deemed markers suitable on semithin sections of plastic-embedded attached oral mucosa. (c) 2008 S. Karger AG, Basel.

An optimized staining technique for the detection of Gram positive and Gram negative bacteria within tissue.

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Becerra, Sandra C; Roy, Daniel C; Sanchez, Carlos J; Christy, Robert J; Burmeister, David M

2016-04-12

Bacterial infections are a common clinical problem in both acute and chronic wounds. With growing concerns over antibiotic resistance, treatment of bacterial infections should only occur after positive diagnosis. Currently, diagnosis is delayed due to lengthy culturing methods which may also fail to identify the presence of bacteria. While newer costly bacterial identification methods are being explored, a simple and inexpensive diagnostic tool would aid in immediate and accurate treatments for bacterial infections. Histologically, hematoxylin and eosin (H&E) and Gram stains have been employed, but are far from optimal when analyzing tissue samples due to non-specific staining. The goal of the current study was to develop a modification of the Gram stain that enhances the contrast between bacteria and host tissue. A modified Gram stain was developed and tested as an alternative to Gram stain that improves the contrast between Gram positive bacteria, Gram negative bacteria and host tissue. Initially, clinically relevant strains of Pseudomonas aeruginosa and Staphylococcus aureus were visualized in vitro and in biopsies of infected, porcine burns using routine Gram stain, and immunohistochemistry techniques involving bacterial strain-specific fluorescent antibodies as validation tools. H&E and Gram stain of serial biopsy sections were then compared to a modification of the Gram stain incorporating a counterstain that highlights collagen found in tissue. The modified Gram stain clearly identified both Gram positive and Gram negative bacteria, and when compared to H&E or Gram stain alone provided excellent contrast between bacteria and non-viable burn eschar. Moreover, when applied to surgical biopsies from patients that underwent burn debridement this technique was able to clearly detect bacterial morphology within host tissue. We describe a modification of the Gram stain that provides improved contrast of Gram positive and Gram negative microorganisms within host

Measurement of cytokine and adhesion molecule expression in synovial tissue by digital image analysis

NARCIS (Netherlands)

Kraan, M. C.; Smith, M. D.; Weedon, H.; Ahern, M. J.; Breedveld, F. C.; Tak, P. P.

2001-01-01

Digital image analysis (DIA) offers the opportunity to quantify the stained area and staining intensity when synovial tissue (ST) is investigated by immunohistochemical analysis. This study aimed at determining the sensitivity of DIA compared with semiquantitative analysis (SQA). Paired ST samples

Automated immunohistochemical method to analyze large areas of the human cortex.

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Abbass, Mohamad; Trought, Kathleen; Long, David; Semechko, Anton; Wong, Albert H C

2018-01-15

There have been inconsistencies in the histological abnormalities found in the cerebral cortex from patients with schizophrenia, bipolar disorder and major depression. Discrepancies in previously published reports may arise from small sample sizes, inconsistent methodology and biased cell counting. We applied automated quantification of neuron density, neuron size and cortical layer thickness in large regions of the cerebral cortex in psychiatric patients. This method accurately segments DAPI positive cells that are also stained with CUX2 and FEZF2. Cortical layer thickness, neuron density and neuron size were automatically computed for each cortical layer in numerous Brodmann areas. We did not find pronounced cytoarchitectural abnormalities in the anterior cingulate cortex or orbitofrontal cortex in patients with schizophrenia, bipolar disorder or major depressive disorder. There were no significant differences in layer thickness measured in immunohistochemically stained slides compared with traditional Nissl stained slides. Automated cell counts were correlated, reliable and consistent with manual counts, while being much less time-consuming. We demonstrate the validity of using a novel automated analysis approach to post-mortem brain tissue. We were able to analyze large cortical areas and quantify specific cell populations using immunohistochemical markers. Future analyses could benefit from efficient automated analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

Lewis x is highly expressed in normal tissues: a comparative immunohistochemical study and literature revision.

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Croce, María V; Isla-Larrain, Marina; Rabassa, Martín E; Demichelis, Sandra; Colussi, Andrea G; Crespo, Marina; Lacunza, Ezequiel; Segal-Eiras, Amada

2007-01-01

An immunohistochemical analysis was employed to determine the expression of carbohydrate antigens associated to mucins in normal epithelia. Tissue samples were obtained as biopsies from normal breast (18), colon (35) and oral cavity mucosa (8). The following carbohydrate epitopes were studied: sialyl-Lewis x, Lewis x, Lewis y, Tn hapten, sialyl-Tn and Thomsen-Friedenreich antigen. Mucins were also studied employing antibodies against MUC1, MUC2, MUC4, MUC5AC, MUC6 and also normal colonic glycolipid. Statistical analysis was performed and Kendall correlations were obtained. Lewis x showed an apical pattern mainly at plasma membrane, although cytoplasmic staining was also found in most samples. TF, Tn and sTn haptens were detected in few specimens, while sLewis x was found in oral mucosa and breast tissue. Also, normal breast expressed MUC1 at a high percentage, whereas MUC4 was observed in a small number of samples. Colon specimens mainly expressed MUC2 and MUC1, while most oral mucosa samples expressed MUC4 and MUC1. A positive correlation between MUC1VNTR and TF epitope (r=0.396) was found in breast samples, while in colon specimens MUC2 and colonic glycolipid versus Lewis x were statistically significantly correlated (r=0.28 and r=0.29, respectively). As a conclusion, a defined carbohydrate epitope expression is not exclusive of normal tissue or a determined localization, and it is possible to assume that different glycoproteins and glycolipids may be carriers of carbohydrate antigens depending on the tissue localization considered.

Immunohistochemical distribution of phosphatidylglucoside using anti-phosphatidylglucoside monoclonal antibody (DIM21)

International Nuclear Information System (INIS)

Kitamura, Yukisato; Okazaki, Toshiro; Nagatsuka, Yasuko; Hirabayashi, Yoshio; Kato, Shinsuke; Hayashi, Kazuhiko

2007-01-01

The immunohistochemical distribution of phosphatidylglucoside (PhGlc) in organs obtained from human autopsy cases was investigated using the DIM21 antibody. Immunohistochemical staining was performed on formaline-fixed, paraffin-embedded sections using the simple stain peroxidase method. The sections were then subjected to antigen retrieval by microwave irradiation in citrate buffer. PhGlc expression was observed in not only the epithelial but also the non-epithelial components of several visceral organs. Squamous and glandular epithelial cells were positive for PhGlc in several organs. The surface areas of the epithelium, particularly the squamous epithelium, were positive. Mesothelial cells were also positive in some organs. Endothelial cells, polymorphonuclear (PMN) cells are positive in several organs. Macrophage is positive in many organs. Epithelial cells of the gallbladder were positive, however, the intrahepatic bile ducts were not positive. In the brain tissue, astroglial cells, the chorioide plexus, the pituitary gland, and ependymal cells were positive. Further investigation is indispensable in order to establish a relationship between cell differentiation and PhGlc expression

Estrogen receptors in human thyroid gland. An immunohistochemical study

International Nuclear Information System (INIS)

Arain, Shaukat A.; Shah, Munawar H.; Jamal, Qamar; Meo, Sultan A.

2003-01-01

The objective of this study is to determine the estrogen receptors (ER) status (present in the nucleous of cell) in the thyroid gland tissues. For this purpose 50 previously diagnosed cases of various thyroid lesions were selected from the Surgical Pathology Records of Pathology Department, Basic Medical Sciences Institute,Jinnah Postgraduate. Medical Center,Karachi,Pakistan between March and August 2000.The staining was performed on formalin fixed paraffin embeded tissues using monoclonal anti-ER anti-body (clone1D5).Out of 50 cases,8 were noduler goiter,9 cases of adenoma 19 papillary carcinoma, 10 follicular and 4 cases were of medullary carcinoma. Surrounding normal tissue was available in 25 (50%) cases, 4 non-neoplastic and 21 neoplastic lesions.Out of 50 cases ,10(20%) and 40(80%) were females, the youngest patient was a 15-year-old female and the eldest patient was a 56-years-old male. Despite the availability of normal thyroid tissue and a wide range of lesions, none of our cases showed the positive staining. In contrary to many earlier reports by immunohistochemical method using monoclonal antibody (clone1D5) on formalin- fixed praffin-embedded thyroid tissues, the ER is not detectable. The effect of Estrogen on thyroid gland may be indirect one. (author)

Optimal iodine staining of cardiac tissue for X-ray computed tomography.

Science.gov (United States)

Butters, Timothy D; Castro, Simon J; Lowe, Tristan; Zhang, Yanmin; Lei, Ming; Withers, Philip J; Zhang, Henggui

2014-01-01

X-ray computed tomography (XCT) has been shown to be an effective imaging technique for a variety of materials. Due to the relatively low differential attenuation of X-rays in biological tissue, a high density contrast agent is often required to obtain optimal contrast. The contrast agent, iodine potassium iodide ([Formula: see text]), has been used in several biological studies to augment the use of XCT scanning. Recently I2KI was used in XCT scans of animal hearts to study cardiac structure and to generate 3D anatomical computer models. However, to date there has been no thorough study into the optimal use of I2KI as a contrast agent in cardiac muscle with respect to the staining times required, which has been shown to impact significantly upon the quality of results. In this study we address this issue by systematically scanning samples at various stages of the staining process. To achieve this, mouse hearts were stained for up to 58 hours and scanned at regular intervals of 6-7 hours throughout this process. Optimal staining was found to depend upon the thickness of the tissue; a simple empirical exponential relationship was derived to allow calculation of the required staining time for cardiac samples of an arbitrary size.

Comparison of Four PD-L1 Immunohistochemical Assays in Lung Cancer.

Science.gov (United States)

Hendry, Shona; Byrne, David J; Wright, Gavin M; Young, Richard J; Sturrock, Sue; Cooper, Wendy A; Fox, Stephen B

2018-03-01

Four different programmed death ligand 1 immunohistochemical assays are approved or in development as companion or complementary diagnostics to different immunotherapeutic agents in lung carcinoma. We sought to determine whether these assays are technically equivalent and whether one antibody can be used on an alternate staining platform. Serial sections of tissue microarrays constructed from 368 cases of resected lung cancer were stained for 22C3 and 28-8 on the Dako Link 48 platform (Dako, Carpinteria, Ca) and for SP142 and SP263 on the Ventana Benchmark Ultra platform (Ventana Medical Systems, Tucson, AZ) strictly as per product insert. A protocol was developed to use the 22C3 antibody on the Ventana Benchmark Ultra platform. Differences in mean tumor cell and immune cell staining were observed between the four assays (p Link 48 platform and the alternate Ventana Benchmark Ultra platform (ICC = 0.921, κ = 0.897). Concordance between the four programmed death ligand 1 immunohistochemical assays when performed and scored as intended show that apart from 28-8 and 22C3, they cannot be used interchangeably in clinical practice. A protocol was successfully developed to use 22C3 on an alternate platform, which may help to overcome some barriers to implementation. Copyright © 2017 International Association for the Study of Lung Cancer. All rights reserved.

Microwave processing of gustatory tissues for immunohistochemistry

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Bond, Amanda; Kinnamon, John C.

2013-01-01

We use immunohistochemistry to study taste cell structure and function as a means to elucidate how taste receptor cells communicate with nerve fibers and adjacent taste cells. This conventional method, however, is time consuming. In the present study we used taste buds from rat circumvallate papillae to compare conventional immunohistochemical tissue processing with microwave processing for the colocalization of several biochemical pathway markers (PLCβ2, syntaxin-1, IP3R3, α-gustducin) and the nuclear stain, Sytox. The results of our study indicate that in microwave versus conventional immunocytochemistry: (1) fixation quality is improved; (2) the amount of time necessary for processing tissue is decreased; (3) antigen retrieval is no longer needed; (4) image quality is superior. In sum, microwave tissue processing of gustatory tissues is faster and superior to conventional immunohistochemical tissue processing for many applications. PMID:23473796

Thick tissue diffusion model with binding to optimize topical staining in fluorescence breast cancer margin imaging

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Xu, Xiaochun; Kang, Soyoung; Navarro-Comes, Eric; Wang, Yu; Liu, Jonathan T. C.; Tichauer, Kenneth M.

2018-03-01

Intraoperative tumor/surgical margin assessment is required to achieve higher tumor resection rate in breast-conserving surgery. Though current histology provides incomparable accuracy in margin assessment, thin tissue sectioning and the limited field of view of microscopy makes histology too time-consuming for intraoperative applications. If thick tissue, wide-field imaging can provide an acceptable assessment of tumor cells at the surface of resected tissues, an intraoperative protocol can be developed to guide the surgery and provide immediate feedback for surgeons. Topical staining of margins with cancer-targeted molecular imaging agents has the potential to provide the sensitivity needed to see microscopic cancer on a wide-field image; however, diffusion and nonspecific retention of imaging agents in thick tissue can significantly diminish tumor contrast with conventional methods. Here, we present a mathematical model to accurately simulate nonspecific retention, binding, and diffusion of imaging agents in thick tissue topical staining to guide and optimize future thick tissue staining and imaging protocol. In order to verify the accuracy and applicability of the model, diffusion profiles of cancer targeted and untargeted (control) nanoparticles at different staining times in A431 tumor xenografts were acquired for model comparison and tuning. The initial findings suggest the existence of nonspecific retention in the tissue, especially at the tissue surface. The simulator can be used to compare the effect of nonspecific retention, receptor binding and diffusion under various conditions (tissue type, imaging agent) and provides optimal staining and imaging protocols for targeted and control imaging agent.

Intramural Ganglion Structures in Esophageal Atresia: A Morphologic and Immunohistochemical Study

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Biagio Zuccarello

2009-01-01

Full Text Available Introduction and Aim. Disorders of esophageal motility causing dysphagia and gastroesophageal reflux are frequent in survivors to esophageal atresia (EA and distal tracheoesophageal fistula (TEF. The aim of the present study was to investigate the histologic and immunohistochemical features in both esophageal atretic segments to further understand the nature of the motor disorders observed in these patients. Material and Methods. Esophageal specimens from 12 newborns with EA/TEF and 5 newborns dead of unrelated causes were examined. The specimens were fixed in 5% buffered formalin, included in paraffin and cut in 5 micron sections that were stained with hematoxilin and eosin (H and E, and immunohistochemical stainings for Actin, S-100 protein, Neurofilament, Neuron-Specific-Enolase, Chromogranin A and Peripherin were evaluated under the microscope. Results. In controls, the distribution of the neural elements was rather homogenous at both levels of the esophagus. In contrast, the atretic segments showed quantitative and qualitative differences between them with sparser nervous tissue in the distal one in comparison with the proximal one and with controls. Conclusions. These results further support the assumption that histomorphological alterations of the muscular and nervous elements within the esophageal wall might contribute to esophageal dysmotility in patients surviving neonatal operations for EA/TEF.

Immunohistochemical quantification of expression of a tight junction protein, claudin-7, in human lung cancer samples using digital image analysis method.

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Lu, Zhe; Liu, Yi; Xu, Junfeng; Yin, Hongping; Yuan, Haiying; Gu, Jinjing; Chen, Yan-Hua; Shi, Liyun; Chen, Dan; Xie, Bin

2018-03-01

Tight junction proteins are correlated with cancer development. As the pivotal proteins in epithelial cells, altered expression and distribution of different claudins have been reported in a wide variety of human malignancies. We have previously reported that claudin-7 was strongly expressed in benign bronchial epithelial cells at the cell-cell junction while expression of claudin-7 was either altered with discontinued weak expression or completely absent in lung cancers. Based on these results, we continued working on the expression pattern of claudin-7 and its relationship with lung cancer development. We herein proposed a new Digital Image Classification, Fragmentation index, Morphological analysis (DICFM) method for differentiating the normal lung tissues and lung cancer tissues based on the claudin-7 immunohistochemical staining. Seventy-seven lung cancer samples were obtained from the Second Affiliated Hospital of Zhejiang University and claudin-7 immunohistochemical staining was performed. Based on C++ and Open Source Computer Vision Library (OpenCV, version 2.4.4), the DICFM processing module was developed. Intensity and fragmentation of claudin-7 expression, as well as the morphological parameters of nuclei were calculated. Evaluation of results was performed using Receiver Operator Characteristic (ROC) analysis. Agreement between these computational results and the results obtained by two pathologists was demonstrated. The intensity of claudin-7 expression was significantly decreased while the fragmentation was significantly increased in the lung cancer tissues compared to the normal lung tissues and the intensity was strongly positively associated with the differentiation of lung cancer cells. Moreover, the perimeters of the nuclei of lung cancer cells were significantly greater than that of the normal lung cells, while the parameters of area and circularity revealed no statistical significance. Taken together, our DICFM approach may be applied as an

Immunohistochemical Assessment of Expression of Centromere Protein—A (CENPA) in Human Invasive Breast Cancer

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Rajput, Ashish B.; Hu, Nianping; Varma, Sonal; Chen, Chien-Hung; Ding, Keyue; Park, Paul C.; Chapman, Judy-Anne W.; SenGupta, Sandip K.; Madarnas, Yolanda; Elliott, Bruce E.; Feilotter, Harriet E.

2011-01-01

Abnormal cell division leading to the gain or loss of entire chromosomes and consequent genetic instability is a hallmark of cancer. Centromere protein –A (CENPA) is a centromere-specific histone-H3-like variant gene involved in regulating chromosome segregation during cell division. CENPA is one of the genes included in some of the commercially available RNA based prognostic assays for breast cancer (BCa)—the 70 gene signature MammaPrint ® and the five gene Molecular Grade Index (MGI SM ). Our aim was to assess the immunohistochemical (IHC) expression of CENPA in normal and malignant breast tissue. Clinically annotated triplicate core tissue microarrays of 63 invasive BCa and 20 normal breast samples were stained with a monoclonal antibody against CENPA and scored for percentage of visibly stained nuclei. Survival analyses with Kaplan–Meier (KM) estimate and Cox proportional hazards regression models were applied to assess the associations between CENPA expression and disease free survival (DFS). Average percentage of nuclei visibly stained with CENPA antibody was significantly higher (p = 0.02) in BCa than normal tissue. The 3-year DFS in tumors over-expressing CENPA (>50% stained nuclei) was 79% compared to 85% in low expression tumors (<50% stained nuclei). On multivariate analysis, IHC expression of CENPA showed weak association with DFS (HR > 60.07; p = 0.06) within our small cohort. To the best of our knowledge, this is the first published report evaluating the implications of increased IHC expression of CENPA in paraffin embedded breast tissue samples. Our finding that increased CENPA expression may be associated with shorter DFS in BCa supports its exploration as a potential prognostic biomarker

Immunohistochemical Assessment of Expression of Centromere Protein—A (CENPA) in Human Invasive Breast Cancer

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Rajput, Ashish B. [Department of Pathology and Molecular Medicine, Queen' s University, Kingston, ON K7L 3N6 (Canada); Hu, Nianping [Cancer Research institute, Queen' s University, Kingston, ON K7L 3N6 (Canada); Varma, Sonal; Chen, Chien-Hung [Department of Pathology and Molecular Medicine, Queen' s University, Kingston, ON K7L 3N6 (Canada); Ding, Keyue [NCIC Clinical Trials Group, Queen' s University, Kingston, ON K7L 3N6 (Canada); Park, Paul C. [Department of Pathology and Molecular Medicine, Queen' s University, Kingston, ON K7L 3N6 (Canada); Chapman, Judy-Anne W. [NCIC Clinical Trials Group, Queen' s University, Kingston, ON K7L 3N6 (Canada); SenGupta, Sandip K. [Department of Pathology and Molecular Medicine, Queen' s University, Kingston, ON K7L 3N6 (Canada); Madarnas, Yolanda [Cancer Research institute, Queen' s University, Kingston, ON K7L 3N6 (Canada); Department of Oncology, Cancer Center of Southeastern Ontario, Kingston, ON K7L 2V7 (Canada); Elliott, Bruce E.; Feilotter, Harriet E., E-mail: feilotth@kgh.kari.net [Department of Pathology and Molecular Medicine, Queen' s University, Kingston, ON K7L 3N6 (Canada)

2011-12-06

Abnormal cell division leading to the gain or loss of entire chromosomes and consequent genetic instability is a hallmark of cancer. Centromere protein –A (CENPA) is a centromere-specific histone-H3-like variant gene involved in regulating chromosome segregation during cell division. CENPA is one of the genes included in some of the commercially available RNA based prognostic assays for breast cancer (BCa)—the 70 gene signature MammaPrint{sup ®} and the five gene Molecular Grade Index (MGI{sup SM}). Our aim was to assess the immunohistochemical (IHC) expression of CENPA in normal and malignant breast tissue. Clinically annotated triplicate core tissue microarrays of 63 invasive BCa and 20 normal breast samples were stained with a monoclonal antibody against CENPA and scored for percentage of visibly stained nuclei. Survival analyses with Kaplan–Meier (KM) estimate and Cox proportional hazards regression models were applied to assess the associations between CENPA expression and disease free survival (DFS). Average percentage of nuclei visibly stained with CENPA antibody was significantly higher (p = 0.02) in BCa than normal tissue. The 3-year DFS in tumors over-expressing CENPA (>50% stained nuclei) was 79% compared to 85% in low expression tumors (<50% stained nuclei). On multivariate analysis, IHC expression of CENPA showed weak association with DFS (HR > 60.07; p = 0.06) within our small cohort. To the best of our knowledge, this is the first published report evaluating the implications of increased IHC expression of CENPA in paraffin embedded breast tissue samples. Our finding that increased CENPA expression may be associated with shorter DFS in BCa supports its exploration as a potential prognostic biomarker.

Functional and immunohistochemical evaluation of porcine neonatal islet-like cell clusters

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Nielsen, T B; Yderstraede, K B; Schrøder, H D

2003-01-01

Porcine neonatal islet-like cell clusters (NICCs) may be an attractive source of insulin-producing tissue for xenotransplantation in type I diabetic patients. We examined the functional and immunohistochemical outcome of the islet grafts in vitro during long-term culture and in vivo after...... transplantation to athymic nude mice. On average we obtained 29,000 NICCs from each pancreas. In a perifusion system, NICCs responded poorly to a glucose challenge alone, but 10 mmol/L arginine elicited a fourfold increase in insulin secretion and 16.7 mmol/L glucose + 10 mmol/L arginine caused a sevenfold...... co-stained for proliferation. However no co-staining was observed between insulin- and glucagon-positive cells or between hormone-and CK-positive cells. Following transplantation of 2000 NICCs under the renal capsule of diabetic nude mice, BG levels were normalized within an average of 13 weeks. Oral...

Experimental infection of octopus vulgaris (Cuvier, 1797) with Photobacterium damsela subsp. piscicida. Immunohistochemical tracking of antigen and tissue responses.

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Bakopoulos, Vasileios; White, Daniella; Valsamidis, Michail-Aggelos; Vasilaki, Feli

2017-03-01

Adult common octopus individuals were intramuscularly infected with Photobacterium damsela subsp. piscicida in order to investigate if this species is sensitive to this common and important fish pathogen. The fate of the bacterial antigens and the tissue responses of Octopus vulgaris were studied employing immunohistochemical techniques. Strong reaction at the site of injection was evident from day 2 post-infection that continued until day 14. Great numbers of hemocytes that were attracted at the site of infection were involved in phagocytosis of bacteria. Very early in the infection, a transition of cells to fibroblasts and an effort to isolate the infection was observed. During the course of the study, very large necrotic cells were seen at the site of infection, whereas during the later stages hemocytes with phagocytosed bacteria were observed in well-defined pockets inside the muscle tissue. None of the internal organs tested for the presence of the bacterium were positive with the exception of the digestive gland where antigen staining was observed which was not associated with hemocyte infiltration. The high doses of bacterial cells used in this experimental infection and the lack of disease signs from Octopus vulgaris suggest that, under normal conditions, octopus is resistant to Photobacterium damsela subsp. piscicida. Copyright © 2017 Elsevier Inc. All rights reserved.

Immunohistochemical study of the expression of cell cycle regulating proteins at different stages of bladder cancer

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Primdahl, Hanne; von der Maase, Hans; Sørensen, Flemming Brandt

2002-01-01

PURPOSE: The cell cycle is known to be deregulated in cancer. We therefore analyzed the expression of the cell cycle related proteins p21, p27, p16, Rb, and L-myc by immunohistochemical staining of bladder tumors.METHODS: The tissue material consisted of bladder tumors from three groups of patients......; group 1, 23 patients with recurrent stage Ta (non-invasive) tumors; group 2, 22 patients presenting at their first admission with T2-4 (muscle invasive) tumors; group 3, 24 patients who experienced disease progression from Ta or T1 (invasive in connective tissue) to a higher stage...

Differential staining of bacteria: gram stain.

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Moyes, Rita B; Reynolds, Jackie; Breakwell, Donald P

2009-11-01

In 1884, Hans Christian Gram, a Danish doctor, developed a differential staining technique that is still the cornerstone of bacterial identification and taxonomic division. This multistep, sequential staining protocol separates bacteria into four groups based on cell morphology and cell wall structure: Gram-positive cocci, Gram-negative cocci, Gram-positive rods, and Gram-negative rods. The Gram stain is useful for assessing bacterial contamination of tissue culture samples or for examining the Gram stain status and morphological features of bacteria isolated from mixed or isolated bacterial cultures. (c) 2009 by John Wiley & Sons, Inc.

An immunohistochemical approach to differentiate hepatic lipidosis from hepatic phospholipidosis in rats.

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Obert, Leslie A; Sobocinski, Gregg P; Bobrowski, Walter F; Metz, Alan L; Rolsma, Mark D; Altrogge, Douglas M; Dunstan, Robert W

2007-08-01

Hepatocellular vacuolation can be a diagnostic challenge since cytoplasmic accumulations of various substances (lipid, water, phospholipids, glycogen, and plasma) can have a similar morphology. Cytoplasmic accumulation of phospholipids following administration of cationic amphiphilic drugs (CAD) can be particularly difficult to differentiate from nonphosphorylated lipid accumulations at the light microscopic level. Histochemical methods (Sudan Black, Oil Red-O, Nile Blue, etc.) can be used to identify both nonphosphorylated and/or phosphorylated lipid accumulations, but these techniques require non-paraffin-embedded tissue and are only moderately sensitive. Thus, electron microscopy is often utilized to achieve a definitive diagnosis based upon the characteristic morphologic features of phospholipid accumulations; however, this is a low throughput and labor intense procedure. In this report, we describe the use of immunohistochemical staining for LAMP-2 (a lysosome-associated protein) and adipophilin (a protein that forms the membrane around non-lysosomal lipid droplets) to differentiate phospholipidosis and lipidosis, respectively in the livers of rats. This staining procedure can be performed on formalin-fixed paraffin embedded tissues, is more sensitive than histochemistry, and easier to perform than ultrastructural evaluation.

Immunohistochemical expression study of proapoptotic BH3-only protein bad in canine nonneoplastic tissues and canine lymphomas.

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Dettwiler, M; Croci, M; Vaughan, L; Guscetti, F

2013-09-01

The BH3-only protein Bad is a proapoptotic Bcl-2 family member that acts as a sensitizer in intrinsic apoptosis by inactivating antiapoptotic members through heterodimer formation. Bad has been shown to contribute to tumorigenesis, including lymphoma formation in humans and mice, through alteration in expression or functional status. Here, its immunohistochemical expression was analyzed in canine nonneoplastic and lymphoma tissues using tissue microarrays. Bad was expressed in the cytoplasm of a wide range of nonneoplastic tissues, especially epithelial cells. Nonneoplastic lymph nodes displayed weak immunostaining in the follicular germinal centers only. Immunoblotting supported these observations but also revealed presence of nonspecific labeling in some organs. Of 81 lymphomas, 29 (35.8%) displayed moderate to strong immunohistochemical Bad labeling, and a significant expression increase was found in lymphomas (especially B cell and double negative) compared to nonneoplastic lymph nodes. These findings warrant further investigations of the functional status, the involvement of partner proteins, and a possible impact of Bad on prognosis in canine lymphoma.

Validation of anti-FXR1 antibodies in the canine species and application to an immunohistochemical study of canine oral melanomas

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Laura Nordio

2017-05-01

Full Text Available FXR1 (Fragile X mental retardation-related protein 1 is a cytoplasmic RNA binding protein, which genetic expression has been related to metastatic potential in human melanoma. The aims of the present study were: the validation of two commercially available clones of polyclonal anti-human FXR1 antibody in dogs; their application to investigate FXR1 expression in a group of canine oral melanomas. Anti-FXR1 antibody was not previously validated in the canine species. Two different commercially available polyclonal anti-FXR1 antibodies (respectively made in goat and in rabbit were used. FXR1 protein in canine serum was identified by western blot after SDS-PAGE, using human serum as control. FXR1 immunohistochemical expression was tested in a series of normal tissues, that are expected to express FXR1, and in 31 cases of oral melanomas. The final immunohistochemical protocol used heat-induced unmasking and overnight incubation. FXR1 protein bands in canine serum were detected by tested antibodies, in a more specific way by the rabbit antibody. FXR1 immunohistochemical staining was positive in all tested organs, with different levels of expression. FXR1 was also expressed in 31/31 tested melanomas, with variable intensity and percentage of positive cells (Figure 1. Equal results were achieved with the two antibodies in 8 cases of melanoma, whereas there were variable differences in 22, and one case stained only with goat antibody. The rabbit antibody gave less background staining. This study validated anti-FXR1 antibodies for use in the canine species. This protein was expressed in various normal tissues, as well as in the tested neoplasms. Significance of different level of expression is undergoing evaluation with further studies.

Giant cell tumor of soft tissues of low malignant potential: A rare diagnosis on fine needle aspiration cytology

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Maithili M Kulkarni

2016-01-01

Full Text Available Primary giant cell tumors of soft tissues (GCT-ST are extremely rare soft tissue tumors, located in both superficial and deep soft tissues. They resemble osseous giant cell tumors morphologically and immunohistochemically. The tumor exhibits strong positive immunoreactivity for cluster of differentiation 68 (CD68 within multinucleated osteoclast-like giant cells and focal staining of mononuclear cells. Case reports describing the cytohistological features of this entity are very few. We report a case of GCT-ST of low malignant potential diagnosed on fine needle aspiration (FNA and confirmed on histological and immunohistochemical studies.

Immunohistochemical analysis of the gingiva with periodontitis of type I plasminogen deficiency compared to gingiva with gingivitis and periodontitis and healthy gingiva.

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Kurtulus Waschulewski, Idil; Gökbuget, Aslan Y; Christiansen, Nina M; Ziegler, Maike; Schuster, Volker; Wahl, Gerhard; Götz, Werner

2016-12-01

Type I plasminogen deficiency (Plgdef) is an uncommon chronic inflammation of mucous membranes. Gingival enlargements usually proceed with progressive periodontal destruction and tooth-loss. Plasmin(ogen)-independent enzymatic mechanisms for fibrin clearance have already been discussed in the literature. Our primary objective was to verify, immunohistochemically, the occurrence of different enzymatic factors involved in tissue breakdown of inflamed compared to healthy gingiva. Secondly, we tried to find out, if these patients have a similar microbiological profile to the patients with known gingivitis and periodontitis. Immunohistochemical analysis of enzymes elastase, plasminogen (plg), cathepsin G, matrix-metalloproteinase (MMP)-3 and MMP-7 and of glycoprotein fibrinogen were performed with gingival tissues from 3 healthy controls, 8 patients with Plgdef and 3 patients with gingivitis and periodontitis. Furthermore, plaque from 5 patients with plasminogen deficiency were also obtained to determine the microbiological profile. Significantly high numbers of elastase positive leukocytes were detected in all samples. Staining for MMP-3 and MMP-7 was seen in samples with gingivitis and periodontitis with a stronger staining in samples with periodontitis by Plgdef. Fibrinogen was detectable in all samples. Staining for plg was stronger in samples with periodontitis than in other samples. Staining for cathepsin G was weak in gingivitis and periodontitis. Subgingival microbial flora showed elevated colony forming units of Prevotella intermedia/nigrescens, Fusobacterium spp., Eikenella corrodens, Porphyromonas gingivalis and viridans streptococci. Strong staining of elastase, MMP-3 and MMP-7 and weak staining of plg in Plgdef samples supports the plasmin(ogen) - independent fibrin clearance. Similar subgingival microbiological flora was observed in periodontitis with Plgdef as in other periodontal diseases. Further investigations should determine the exact pathomechanism

Using cell nuclei features to detect colon cancer tissue in hematoxylin and eosin stained slides.

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Jørgensen, Alex Skovsbo; Rasmussen, Anders Munk; Andersen, Niels Kristian Mäkinen; Andersen, Simon Kragh; Emborg, Jonas; Røge, Rasmus; Østergaard, Lasse Riis

2017-08-01

Currently, diagnosis of colon cancer is based on manual examination of histopathological images by a pathologist. This can be time consuming and interpretation of the images is subject to inter- and intra-observer variability. This may be improved by introducing a computer-aided diagnosis (CAD) system for automatic detection of cancer tissue within whole slide hematoxylin and eosin (H&E) stains. Cancer disrupts the normal control mechanisms of cell proliferation and differentiation, affecting the structure and appearance of the cells. Therefore, extracting features from segmented cell nuclei structures may provide useful information to detect cancer tissue. A framework for automatic classification of regions of interest (ROI) containing either benign or cancerous colon tissue extracted from whole slide H&E stained images using cell nuclei features was proposed. A total of 1,596 ROI's were extracted from 87 whole slide H&E stains (44 benign and 43 cancer). A cell nuclei segmentation algorithm consisting of color deconvolution, k-means clustering, local adaptive thresholding, and cell separation was performed within the ROI's to extract cell nuclei features. From the segmented cell nuclei structures a total of 750 texture and intensity-based features were extracted for classification of the ROI's. The nine most discriminative cell nuclei features were used in a random forest classifier to determine if the ROI's contained benign or cancer tissue. The ROI classification obtained an area under the curve (AUC) of 0.96, sensitivity of 0.88, specificity of 0.92, and accuracy of 0.91 using an optimized threshold. The developed framework showed promising results in using cell nuclei features to classify ROIs into containing benign or cancer tissue in H&E stained tissue samples. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

Immunohistochemical evaluation of fibrovascular and cellular pre-iridal membranes in dogs.

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Bauer, Bianca S; Sandmeyer, Lynne S; Hall, Riley B; Grahn, Bruce H

2012-03-01

Histologically, two morphologically distinct types of pre-iridal membranes appear to occur in diseased canine globes: fibrovascular and cellular. Cellular pre-iridal membranes of corneal endothelial origin exist in iridocorneal endothelial (ICE) syndrome in humans and arise through metaplastic transformation of corneal endothelial cells into epithelial-like cells.(1) The purpose of this study was to (i) evaluate immunohistochemical staining of these two types of membranes in diseased canine globes, (ii) determine whether endothelial cell metaplasia or iridal vascular budding plays a role in cellular membrane formation and (iii) compare the primary histopathologic diagnosis between the two groups. Hematoxylin and eosin (H&E)-stained slides of 28 enucleated canine specimens with pre-iridal membranes were randomly selected and examined with light microscopy. The globes were divided into two groups based on the appearance of the membrane: fibrovascular or cellular, and the histopathologic diagnoses were recorded. Immunohistochemical staining for vimentin, cytokeratin AE1/AE3, and Von Willebrand's factor (Factor VIII) was completed on the slides of each globe. The histopathologic diagnoses were compared between the two groups. The fibrovascular and cellular membranes stained positive for vimentin and negative for cytokeratin AE1/AE3. All fibrovascular membranes stained positive for Factor VIII compared with the cellular membranes which stained negative. In the cellular membrane group, primary glaucoma was a common histologic diagnosis. Immunohistochemical evaluation in this study does not support the hypothesis of metaplastic transformation of endothelial cells into epithelial-like cells in the canine globes with cellular membranes. The cellular membranes in this study do not represent a canine version of ICE syndrome and are not of vascular endothelial origin. © 2012 American College of Veterinary Ophthalmologists.

Immunohistochemical detection of XIAP in melanoma.

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Emanuel, Patrick O M; Phelps, Robert G; Mudgil, Adarsh; Shafir, Michail; Burstein, David E

2008-03-01

The X-linked inhibitor of apoptosis protein (XIAP) is the most potent of the inhibitor of apoptosis family of eight proteins. High levels of XIAP have been found in melanoma cell lines and are believed to play a role in therapeutic resistance in a number of malignancies. XIAP expression has not been investigated in clinically obtained melanoma tissue samples, nor have studies attempted to correlate XIAP expression with prognostic variables or clinical aggressiveness of melanomas. Sixty-seven patients with primary cutaneous malignant melanoma for whom clinical follow up was available were identified from the records of the Mount Sinai Hospital, comprising 37 thin melanomas (Breslow thickness 1.0 mm). Archival paraffin sections from primary lesions and corresponding metastases were stained with monoclonal anti-XIAP antibody using routine immunohistochemical methods. Six benign intradermal nevi and four in situ melanomas were XIAP negative. 9 of 37 thin melanomas (24%) were XIAP positive. In contrast, 21 of 30 (73%) thick melanomas were XIAP positive, including 3 of 4 ulcerated melanomas that were strongly positive. Over a follow-up period ranging from 6 months to 6 years, 23 melanomas metastasized (22 thick, 1 thin). In total, XIAP was immunohistochemically detected in 17 of 23 metastases (74%). Metastasis occurred in 1 of 9 XIAP-positive thin melanomas; 0 of 28 XIAP-negative thin melanomas; 17 of 22 XIAP-positive thick melanomas, and 5 of 8 XIAP-negative thick melanomas (63%). XIAP is immunohistochemically detectable nearly three times more frequently in thick compared with thin melanomas. These results suggest that XIAP elevation may be correlated with increasing melanoma thickness and tumor progression.

Accelerated staining technique using kitchen microwave oven

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Archana Mukunda

2015-01-01

Full Text Available Introduction: Histopathological diagnosis of specimens is greatly dependent on good sample preparation and staining. Both of these processes is governed by diffusion of fluids and dyes in and out of the tissue, which is the key to staining. Diffusion of fluids can be accelerated by the application of heat that reduces the time of staining from hours to the minute. We modified an inexpensive model of kitchen microwave oven for staining. This study is an attempt to compare the reliability of this modified technique against the tested technique of routine staining so as to establish the kitchen microwave oven as a valuable diagnostic tool. Materials and Methods: Sixty different tissue blocks were used to prepare 20 pairs of slides for 4 different stains namely hematoxylin and eosin, Van Gieson′s, 0.1% toluidine blue and periodic acid-Schiff. From each tissue block, two bits of tissues were mounted on two different slides. One slide was stained routinely, and the other stained inside a microwave. A pathologist evaluated the stained slides and the results so obtained were analyzed statistically. Results: Microwave staining considerably cut down the staining time from hours to seconds. Microwave staining showed no loss of cellular and nuclear details, uniform-staining characteristics and was of excellent quality. Interpretation and Conclusion: The cellular details, nuclear details and staining characteristics of microwave stained tissues were better than or equal to the routine stained tissue. The overall quality of microwave-stained sections was found to be better than the routine stained tissue in majority of cases.

Electrochemical removal of metallic implants from Technovit 9100 New embedded hard and soft tissues prior to histological sectioning.

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Willbold, Elmar; Reebmann, Mattias; Jeffries, Richard; Witte, Frank

2013-11-01

Solid metallic implants in soft or hard tissues are serious challenges for histological processing. However, metallic implants are more frequently used in e.g. cardiovascular or orthopaedic therapies. Before clinical use, these devices need to be tested thoroughly in a biological environment and histological analysis of their biocompatibility is a major requirement. To allow the histological analysis of metallic implants in tissues especially in calcified hard tissues, we describe a method for embedding these tissues in the resin Technovit 9100 New and removing the metallic implants by electrochemical dissolution. With the combination of these two processes, we are able to achieve 5 μm thick sections from soft or hard tissues with a superior preservation of tissue architecture and especially the implant-tissue interface. These sections can be stained by classical stainings, immunohistochemical and enzymehistochemical as well as DNA-based staining methods.

Immunohistochemical testing for Helicobacter Pylori existence in neoplasms of the colon

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Yasar Nurgul

2008-08-01

Full Text Available Abstract Background Helicobacter pylori is a common pathogen, and its prevalence varies with socioeconomic conditions (10–80%. It has recently been recognized as a class I carcinogen in relation to gastric cancer. The aim of this study was to investigate the presence of Helicobacter pylori in neoplasms of the colon by immunohistochemical methods. Methods The polypectomy materials of 51 patients (19 male and 32 female who had undergone colonoscopic polypectomy were retrieved for retrospective examination. The endoscopic size and colonic localization of the polyps were recorded. Hematoxylin and eosin stains were evaluated according to histological type and grade of dysplasia. Biopsy stains were immunohistochemically treated with Helicobacter pylori antibodies by the streptavidine-biotin immunoperoxidase technique. Helicobacter pylori staining in the gastric mucosa was used as the control for the immunohistochemical method. Specimens were classified according to the presence of Helicobacter pylori under an optical microscope, and Helicobacter pylori positive specimens were stratified according to the respective staining pattern. Results Mean age was 61.88 ± 10.62 (40–82 years. Polyp sizes were 1.45 ± 0.92 (1–4 cm; and 25.5% of polyps were localized in the right colon, 68.6% in the left colon and 5.9% in the transverse colon. Presence of Helicobacter pylori was not correlated with localization (p > 0.05 or size of the polyps (p > 0.05. Eleven (21.6% of all specimens included in the study were Helicobacter pylori positive by immunohistochemical methods. Of the Helicobacter pylori positive specimens, the staining pattern was diffuse: Equivocal in 90.9%, nonspecific with a finely granular type concentrated on the luminal surface in 90.9%, dot-like granular in 54.5%, and spiral in 9.1%. Of the tubular polyps, 17.9% were H. pylori positive, and the staining pattern was equivocal in 100%, luminal in 85.7%, and dot-like granular in 57.1%. Of the

Expression of CA125 in Tissue and Serum of Uterine Serous Carcinoma Patients

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Joseph Menczer

2015-09-01

Conclusion: The higher proportion of USC patients with positive CA125 immunohistochemical tissue staining than with elevated serum CA125 levels indicates the presence of some mechanism that prevents the access of CA125 into the circulation. [J Interdiscipl Histopathol 2015; 3(3.000: 102-104

Combined in situ zymography, immunofluorescence, and staining of iron oxide particles in paraffin-embedded, zinc-fixed tissue sections.

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Haeckel, Akvile; Schoenzart, Lena; Appler, Franziska; Schnorr, Joerg; Taupitz, Matthias; Hamm, Bernd; Schellenberger, Eyk

2012-01-01

Superparamagnetic iron oxide particles are used as potent contrast agents in magnetic resonance imaging. In histology, these particles are frequently visualized by Prussian blue iron staining of aldehyde-fixed, paraffin-embedded tissues. Recently, zinc salt-based fixative was shown to preserve enzyme activity in paraffin-embedded tissues. In this study, we demonstrate that zinc fixation allows combining in situ zymography with fluorescence immunohistochemistry (IHC) and iron staining for advanced biologic investigation of iron oxide particle accumulation. Very small iron oxide particles, developed for magnetic resonance angiography, were applied intravenously to BALB/c nude mice. After 3 hours, spleens were explanted and subjected to zinc fixation and paraffin embedding. Cut tissue sections were further processed to in situ zymography, IHC, and Prussian blue staining procedures. The combination of in situ zymography as well as IHC with subsequent Prussian blue iron staining on zinc-fixed paraffin-embedded tissues resulted in excellent histologic images of enzyme activity, protease distribution, and iron oxide particle accumulation. The combination of all three stains on a single section allowed direct comparison with only moderate degradation of fluorescein isothiocyanate-labeled substrate. This protocol is useful for investigating the biologic environment of accumulating iron oxide particles, with excellent preservation of morphology.

Comparison of special stains for keratin with routine hematoxylin and eosin stain.

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Rao, Roopa S; Patil, Shankargouda; Majumdar, Barnali; Oswal, Rakesh G

2015-03-01

Keratins are the most abundant proteins and are characteristic findings in many epithelial pathologies, making it a diagnostically important marker, both histopathologically and immunohistochemically. Since, immunohistochemistry is an expensive diagnostic tool, special stains to detect the degree of keratinization could serve as a faster and economic option. The aim of the present study was to compare the efficacy of special stains for keratin with standard hematoxylin and eosin stain (H and E). Objectives include: (i) To subject the diagnosed cases of keratin disorders to the selected special stains: Ayoub-shklar method, Dane-Herman method, Alcian blue -periodic acid Schiff 's (PAS), rapid papanicolaou (PAP) and Gram's stain. (ii) To compare the staining specificity and staining intensity of special stains with respect to routine hematoxylin and eosin (H and E) stain. (iii) To compare the efficacy of special stains to routine H and E stain in identification of the type of keratin present in the selected cases. A total of 80 cases of known pathology for keratin were retrieved from the department archive, which included 10 each of normal gingiva, hyperkeratosis, squamous papilloma, verrucous hyperplasia, verrucous carcinoma, well-differentiated squamous cell carcinoma, orthokeratinized odontogenic cyst and keratocystic odontogenic tumors. Six sections of 4 µ each from the paraffin blocks were made, stained with H and E and the special stains and these were evaluated by 2 pathologists based on the modified scoring criteria from Rahma Al-Maaini and Philip Bryant 2008. The results were tabulated using Chi square and kappa statistics. The statistical values for identification of the type of keratinization was insignificant showing that ortho and parakeratinized epithelia could be correctly identified by both H and E as well as all the special stains. Furthermore, all the special stains showed a positive result and statistical significance (P < 0.001) with respect to

Immunohistochemical Expression of TGF-β1 and Osteonectin in engineered and Ca(OH2-repaired human pulp tissues

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Luiz Alexandre CHISINI

Full Text Available Abstract The aim of the present study was to evaluate the expression of transforming growth factor-β1 (TGF-β1 and osteonectin (ON in pulp-like tissues developed by tissue engineering and to compare it with the expression of these proteins in pulps treated with Ca(OH2 therapy. Tooth slices were obtained from non-carious human third molars under sterile procedures. The residual periodontal and pulp soft tissues were removed. Empty pulp spaces of the tooth slice were filled with sodium chloride particles (250–425 µm. PLLA solubilized in 5% chloroform was applied over the salt particles. The tooth slice/scaffold (TS/S set was stored overnight and then rinsed thoroughly to wash out the salt. Scaffolds were previously sterilized with ethanol (100–70° and washed with phosphate-buffered saline (PBS. TS/S was treated with 10% EDTA and seeded with dental pulp stem cells (DPSC. Then, TS/S was implanted into the dorsum of immunodeficient mice for 28 days. Human third molars previously treated with Ca(OH2 for 90 days were also evaluated. Samples were prepared and submitted to histological and immunohistochemical (with anti-TGF-β1, 1:100 and anti-ON, 1:350 analyses. After 28 days, TS/S showed morphological characteristics similar to those observed in dental pulp treated with Ca(OH2. Ca(OH2-treated pulps showed the usual repaired pulp characteristics. In TS/S, newly formed tissues and pre-dentin was colored, which elucidated the expression of TGF-β1 and ON. Immunohistochemistry staining of Ca(OH2-treated pulps showed the same expression patterns. The extracellular matrix displayed a fibrillar pattern under both conditions. Regenerative events in the pulp seem to follow a similar pattern of TGF-β1 and ON expression as the repair processes.

Discriminative staining methods for the nervous system: luxol fast blue--periodic acid-Schiff--hematoxylin triple stain and subsidiary staining methods.

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Goto, N

1987-09-01

This paper describes a new series of staining methods which can discriminatively demonstrate every structure of the nervous system, including axons and capillaries, in animal and human materials. Methods described in this paper consist of one primary stain, luxol fast blue-periodic acid Schiff-hematoxylin (LPH) and six different subsidiary staining methods. The LPH triple stain can precisely differentiate the following structures: neurons (Nissl bodies, cytoplasm, nuclear membrane and nucleolus), various kinds of nuclei (glia, ependyma, endothelium, leucocyte, connective tissue, etc.), myelin sheaths, neuronal processes (axons and dendrites), reacted glial cell bodies (protoplasmic astrocytes, foamy cells, etc.), blood vessels (arteries, veins and capillaries), meninges, intervening connective tissue, erythrocytes, lipofuscin granules, amyloid bodies, and others. Subsidiary staining methods are also described briefly. Applications are discussed in the context of staining technology and neuromorphological research.

Comparative evaluation of eosinophils in normal mucosa, dysplastic mucosa and oral squamous cell carcinoma with hematoxylin-eosin, Congo red, and EMR1 immunohistochemical staining techniques.

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Kargahi, Neda; Razavi, Sayyed Mohammad; Deyhimi, Parviz; Homayouni, Solmaz

2015-01-01

Oral squamous cell carcinoma is the most common malignant lesion of the oral cavity, and it involves various molecular mechanisms. The development of oral squamous cell carcinoma is influenced by the host immune cells, such as eosinophils. The present study was conducted to compare the presence of eosinophils in normal mucosa, dysplastic mucosa, and oral squamous cell carcinoma by -hematoxylin- eosin staining, Congo red staining, and epidermal growth factor-like (EGF-like) module containing a mucin-like hormone receptor1 (EMR1) immunohistochemical marker. In this cross-sectional study, 60 paraffinized samples were selected, consisting of 20 normal mucosae, 20 dysplastic mucosae, and 20 squamous cell carcinoma samples. After confirmation of the diagnosis, the mean number of eosinophils was evaluated by hematoxylin-eosin, Congo red, and immunohystochemical staining techniques. The data were analyzed by SPSS-10 software using the Kruskal-Wallis and Friedman tests. The results showed that the number of eosinophils in dysplastic mucosa was significantly higher than the number in normal mucosa, and the number of eosinophils in squamous cell carcinoma was significantly higher than the number in dysplastic mucosa in all staining techniques (p<0.001). Moreover, the comparison of staining techniques showed a significantly higher number of eosinophils in EMR1immunohistochemicalmarker than were observed when Congo red and hematoxylin - eosin (H&E) staining techniques were used (p<0.001). It can be argued that eosinophil contributes to the identification of lesions that have a higher potential of malignant transformation. Moreover, eosinophil can be suggested as an indicator in the differentiation of oral lesions in cases with borderline diagnosis and in targeted molecular therapy.

Background staining of visualization systems in immunohistochemistry: comparison of the Avidin-Biotin Complex system and the EnVision+ system.

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Vosse, Bettine A H; Seelentag, Walter; Bachmann, Astrid; Bosman, Fred T; Yan, Pu

2007-03-01

The aim of this study was to evaluate specific immunostaining and background staining in formalin-fixed, paraffin-embedded human tissues with the 2 most frequently used immunohistochemical detection systems, Avidin-Biotin-Peroxidase (ABC) and EnVision+. A series of fixed tissues, including breast, colon, kidney, larynx, liver, lung, ovary, pancreas, prostate, stomach, and tonsil, was used in the study. Three monoclonal antibodies, 1 against a nuclear antigen (Ki-67), 1 against a cytoplasmic antigen (cytokeratin), and 1 against a cytoplasmic and membrane-associated antigen and a polyclonal antibody against a nuclear and cytoplasmic antigen (S-100) were selected for these studies. When the ABC system was applied, immunostaining was performed with and without blocking of endogenous avidin-binding activity. The intensity of specific immunostaining and the percentage of stained cells were comparable for the 2 detection systems. The use of ABC caused widespread cytoplasmic and rare nuclear background staining in a variety of normal and tumor cells. A very strong background staining was observed in colon, gastric mucosa, liver, and kidney. Blocking avidin-binding capacity reduced background staining, but complete blocking was difficult to attain. With the EnVision+ system no background staining occurred. Given the efficiency of the detection, equal for both systems or higher with EnVision+, and the significant background problem with ABC, we advocate the routine use of the EnVision+ system.

Duchenne and Becker Muscular Dystrophy: Contribution of a Molecular and Immunohistochemical Analysis in Diagnosis in Morocco

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Hanane Bellayou

2009-01-01

Full Text Available Duchenne muscular dystrophy (DMD and Becker muscular dystrophy (BMD are X-linked recessive disorders caused by mutations of the DMD gene located at Xp21. In DMD patients, dystrophin is virtually absent; whereas BMD patients have 10% to 40% of the normal amount. Deletions in the dystrophin gene represent 65% of mutations in DMD/BMD patients. To explain the contribution of immunohistochemical and genetic analysis in the diagnosis of these dystrophies, we present 10 cases of DMD/BMD with particular features. We have analyzed the patients with immunohistochemical staining and PCR multiplex to screen for exons deletions. Determination of the quantity and distribution of dystrophin by immunohistochemical staining can confirm the presence of dystrophinopathy and allows differentiation between DMD and BMD, but dystrophin staining is not always conclusive in BMD. Therefore, only identification involved mutation by genetic analysis can establish a correct diagnosis.

Uterine Carcinosarcomas: Clinical, Histopathologic and Immunohistochemical Characteristics.

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Chen, Xiaowei; Arend, Rebecca; Hamele-Bena, Diane; Tergas, Ana I; Hawver, Melanie; Tong, Guo-Xia; Wright, Thomas C; Wright, Jason D

2017-09-01

Carcinosarcomas (malignant mixed Müllerian tumors or MMMT) are rare malignant tumors in the female genital tract composed of both malignant epithelial and malignant mesenchymal components. They comprise 1995 and 2011 were retrieved from the gynecologic pathology files at Columbia University Medical Center. Representative tissue blocks containing both epithelial and mesenchymal components were selected from each case for histologic and immunohistochemical studies. Clinical data from each case were retrieved. The epithelial component was poorly differentiated adenocarcinoma in the majority (80.7%) of cases; in 17.7%, the carcinoma was moderately differentiated, and in only 1.6% the carcinoma was well differentiated. 53% of the tumors had homologous stromal elements and 47% displayed heterologous stromal elements. Immunohistochemical study revealed almost equal staining in both epithelial and mesenchymal components of carcinosarcomas for p16 and p53. PAX8 positivity was noted in 73% of epithelial components, but only 13% of stromal components, and PAX8 stromal positivity was never seen in the absence of PAX8 epithelial positivity. Expression of p16, p53, and PAX8 in both malignant components lends support to the monoclonal theory of uterine carcinosarcoma tumorigenesis. The roles of these tumor markers in the diagnosis and pathogenesis of this tumor and associations between clinical characteristics, tumor pathologic features, and prognosis are discussed.

Antibody Selection for Cancer Target Validation of FSH-Receptor in Immunohistochemical Settings

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Nina Moeker

2017-10-01

Full Text Available Background: The follicle-stimulating hormone (FSH-receptor (FSHR has been reported to be an attractive target for antibody therapy in human cancer. However, divergent immunohistochemical (IHC findings have been reported for FSHR expression in tumor tissues, which could be due to the specificity of the antibodies used. Methods: Three frequently used antibodies (sc-7798, sc-13935, and FSHR323 were validated for their suitability in an immunohistochemical study for FSHR expression in different tissues. As quality control, two potential therapeutic anti-hFSHR Ylanthia® antibodies (Y010913, Y010916 were used. The specificity criteria for selection of antibodies were binding to native hFSHR of different sources, and no binding to non-related proteins. The ability of antibodies to stain the paraffin-embedded Flp-In Chinese hamster ovary (CHO/FSHR cells was tested after application of different epitope retrieval methods. Results: From the five tested anti-hFSHR antibodies, only Y010913, Y010916, and FSHR323 showed specific binding to native, cell-presented hFSHR. Since Ylanthia® antibodies were selected to specifically recognize native FSHR, as required for a potential therapeutic antibody candidate, FSHR323 was the only antibody to detect the receptor in IHC/histochemical settings on transfected cells, and at markedly lower, physiological concentrations (ex., in Sertoli cells of human testes. The pattern of FSH323 staining noticed for ovarian, prostatic, and renal adenocarcinomas indicated that FSHR was expressed mainly in the peripheral tumor blood vessels. Conclusion: Of all published IHC antibodies tested, only antibody FSHR323 proved suitable for target validation of hFSHR in an IHC setting for cancer. Our studies could not confirm the previously reported FSHR overexpression in ovarian and prostate cancer cells. Instead, specific overexpression in peripheral tumor blood vessels could be confirmed after thorough validation of the antibodies used.

Immunohistochemical Analysis Using Antipodocalyxin Monoclonal Antibody PcMab-47 Demonstrates Podocalyxin Expression in Oral Squamous Cell Carcinomas.

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Itai, Shunsuke; Yamada, Shinji; Kaneko, Mika K; Harada, Hiroyuki; Kato, Yukinari

2017-10-01

Podocalyxin is a CD34-related type I transmembrane protein that is highly glycosylated with N-glycan, O-glycan, and keratan sulfate. Podocalyxin was originally found in the podocytes of rat kidney and is reportedly expressed in many types of tumors, including brain tumors, colorectal cancers, and breast cancers. Overexpression of podocalyxin is an independent predictor of progression, metastasis, and poor outcome. We recently immunized mice with recombinant human podocalyxin, which was produced using LN229 glioblastoma cells, and produced a novel antipodocalyxin monoclonal antibody (mAb), PcMab-47, which reacts with endogenous podocalyxin-expressing cancer cell lines and normal cell lines independent of glycosylation in Western blot, flow cytometry, and immunohistochemical analyses. In this study, we performed immunohistochemical analysis against oral cancers using PcMab-47. PcMab-47-stained oral squamous cell carcinoma cells in a cytoplasmic pattern and detected 26/38 (68.4%) of oral squamous cell carcinoma cells on tissue microarrays. These results indicate that PcMab-47 is useful in detecting podocalyxin of oral cancers for immunohistochemical analysis.

Angiomyolipoma of the kidney: Clinico pathological and immunohistochemical study

International Nuclear Information System (INIS)

Esheba, Gh.E.; Esheba, N.E.

2013-01-01

Overview: Although angiomyolipoma (AML) is a relatively rare entity, it is the most common benign mesenchymal neoplasm of the kidney. The aim of this study: To highlight the clinicopathological characteristics of AML and to assess the role of Human Melanoma Black-45 (HMB-45), Melan-A, smooth muscle actin (SMA), S-100 and cytokeratin in its diagnosis. Materials and methods: The study included 15 cases of AML. Clinical and radiological data were retrieved from the archival files and all cases were subjected to a histopathological evaluation as well as immunohistochemical staining for HMB-45, Melan-A, SMA, S-100, and cytokeratin. Results: AML was more common in females (female:male = 4:1), the mean age was 53.9 ± 6.45 years. 60% of patients were symptomatic while the remaining 40% were asymptomatic. A statistically significant relationship was found between size of the tumor and the presence of the symptoms (P = 0.02). Patients with tumor size less than 4 cm were asymptomatic, while those with tumor size larger than 4 cm had different symptoms. Thirteen cases were classic AML, while 2 cases were epithelioid AML. Classic AML demonstrated admixture of fatty tissue, thick-walled blood vessels, and smooth muscle, while epithelioid AML was composed mainly of epithelioid cells and contained no fat. HMB-45 was positive in all cases of AML (100%), Melan-A was positive in 13/15 (87%) while SMA was positive in 11/15 (73%) of AML with variable staining intensity. All cases of AML were negative for S-100 and cytokeratin. Conclusion: AMLs have characteristic clinicopathological and immunohistochemical features and their recognition is crucial for proper diagnosis and treatment

Metallothionein expression in placental tissue in Menkes' disease

DEFF Research Database (Denmark)

Hærslev, T.; Krag Jacobsen, G.; Horn, N.

1995-01-01

. The avidin-biotin-complex (ABC)-technique was used. The copper content was measured by neutron activation analysis (NAA). In all placental tissue sections positive MT immunostaining appeared only in the trophoblast and only in proliferating cells. In placental tissue sections obtained from foetuses...... and children affected by Menkes' disease an additional MT immunostaining appeared in the Hofbauer cells of the chorionic villi. This staining was associated with an increased content of copper as measured by NAA. We conclude that the immunohistochemical demonstration of MT reflects the copper content and may...

Immunohistochemical Detection of a Unique Protein within Cells of Snakes Having Inclusion Body Disease, a World-Wide Disease Seen in Members of the Families Boidae and Pythonidae

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Chang, Li-Wen; Fu, Ann; Wozniak, Edward; Chow, Marjorie; Duke, Diane G.; Green, Linda; Kelley, Karen; Hernandez, Jorge A.; Jacobson, Elliott R.

2013-01-01

Inclusion body disease (IBD) is a worldwide disease in captive boa constrictors (boa constrictor) and occasionally in other snakes of the families Boidae and Pythonidae. The exact causative agent(s) and pathogenesis are not yet fully understood. Currently, diagnosis of IBD is based on the light microscopic identification of eosinophilic intracytoplasmic inclusion bodies in hematoxylin and eosin stained tissues or blood smears. An antigenically unique 68 KDa protein was identified within the IBD inclusion bodies, called IBD protein. A validated immuno-based ante-mortem diagnostic test is needed for screening snakes that are at risk of having IBD. In this study, despite difficulties in solubilizing semi-purified inclusion bodies, utilizing hybridoma technology a mouse anti-IBD protein monoclonal antibody (MAB) was produced. The antigenic specificity of the antibody was confirmed and validated by western blots, enzyme-linked immunosorbent assay, immuno-transmission electron microscopy, and immunohistochemical staining. Paraffin embedded tissues of IBD positive and negative boa constrictors (n=94) collected from 1990 to 2011 were tested with immunohistochemical staining. In boa constrictors, the anti-IBDP MAB had a sensitivity of 83% and specificity of 100% in detecting IBD. The antibody also cross-reacted with IBD inclusion bodies in carpet pythons (Morelia spilota) and a ball python (python regius). This validated antibody can serve as a tool for the development of ante-mortem immunodiagnostic tests for IBD. PMID:24340066

Immunohistochemical detection of a unique protein within cells of snakes having inclusion body disease, a world-wide disease seen in members of the families Boidae and Pythonidae.

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Li-Wen Chang

Full Text Available Inclusion body disease (IBD is a worldwide disease in captive boa constrictors (boa constrictor and occasionally in other snakes of the families Boidae and Pythonidae. The exact causative agent(s and pathogenesis are not yet fully understood. Currently, diagnosis of IBD is based on the light microscopic identification of eosinophilic intracytoplasmic inclusion bodies in hematoxylin and eosin stained tissues or blood smears. An antigenically unique 68 KDa protein was identified within the IBD inclusion bodies, called IBD protein. A validated immuno-based ante-mortem diagnostic test is needed for screening snakes that are at risk of having IBD. In this study, despite difficulties in solubilizing semi-purified inclusion bodies, utilizing hybridoma technology a mouse anti-IBD protein monoclonal antibody (MAB was produced. The antigenic specificity of the antibody was confirmed and validated by western blots, enzyme-linked immunosorbent assay, immuno-transmission electron microscopy, and immunohistochemical staining. Paraffin embedded tissues of IBD positive and negative boa constrictors (n=94 collected from 1990 to 2011 were tested with immunohistochemical staining. In boa constrictors, the anti-IBDP MAB had a sensitivity of 83% and specificity of 100% in detecting IBD. The antibody also cross-reacted with IBD inclusion bodies in carpet pythons (Morelia spilota and a ball python (python regius. This validated antibody can serve as a tool for the development of ante-mortem immunodiagnostic tests for IBD.

Immunohistochemical detection of Her-2/neu overexpression in ...

African Journals Online (AJOL)

Materials and Methods: Immunohistochemical staining for Her-2/neu was performed on 10% formalin-fixed, paraffinembedded primary carcinoma of the breast from 83 patients, between 2003 and 2007 using anti-Her-2/neu rabbit polyclonal antibody (DakoCytomation, CA, USA) and reactivity detected by an avidin-biotin ...

YAP expression in normal and neoplastic breast tissue: an immunohistochemical study.

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Jaramillo-Rodríguez, Yolanda; Cerda-Flores, Ricardo M; Ruiz-Ramos, Ruben; López-Márquez, Francisco C; Calderón-Garcidueñas, Ana Laura

2014-04-01

Yes-associated protein (YAP) is a transcriptional factor involved in normal cell proliferation, apoptosis and carcinogenesis; however, its contribution to breast cancer (BC) is still controversial. We undertook this study to compare the expression of YAP by immunohistochemistry (IHC) in normal breast tissue of women without breast cancer (BC) (controls), non-neoplastic breast tissue in women with cancer (internal controls) and in four different subtypes of invasive ductal carcinoma. There were 17 controls and 105 tumor cases (53 luminal A, 15 luminal B, 20 overexpression of HER2 and 17 triple negative cases) studied by IHC. Statistical analysis included χ(2) for linear trend (Extended Mantel-Haenszel). There were 40% of internal controls that showed expression of YAP in myoepithelial cells, whereas in controls expression was 100%. In controls, 3/17 (17.6%) showed cytoplasmic staining in luminal cells. There was a significant difference in nuclear expression between the ductal BC subtypes. Luminal A had 4% of positive cases with <10% of cells affected in each case; in contrast, there were 17-20% of positive cases in the other groups with 50% or more of stained cells. YAP expression in stromal cells was not observed in controls or in triple-negative cases, and luminal B pattern had the highest YAP nuclear expression (20%). YAP showed decreased expression in tumor cells compared with normal breast tissue. These findings are consistent with a role of YAP as a suppressor gene in BC and show differences in YAP expression in different patterns of ductal BC. Copyright © 2014 IMSS. Published by Elsevier Inc. All rights reserved.

The Luna stain, an improved selective stain for detection of microsporidian spores in histologic sections

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Peterson, Tracy S.; Spitsbergen, Jan M.; Feist, Stephen W.; Kent, Michael L.

2014-01-01

Microsporidia in histologic sections are most often diagnosed by observing spores in host tissues. Spores are easy to identify if they occur in large aggregates or xenomas when sections are stained with hematoxylin and eosin (H&E). However, individual spores are not frequently detected in host tissues with conventional H&E staining, particularly if spores are scattered within the tissues, areas of inflammation or small spores in nuclei (i.e., Nucleospora salmonis). Hence, a variety of selective stains that enhance visualization of spores are recommended. We discovered that the Luna stain, used to highlight eosinophils, red blood cells and chitin in arthropods and other invertebrates, also stains spores of Pseudoloma neurophilia. We compared this stain to the Gram, Fite’s acid fast, Giemsa, and H&E stains on eight aquatic microsporidian organisms that were readily available in our two laboratories: Loma salmonae, Glugea anomala, Pseudoloma neurophilia, Pleistophora hyphessobryconis, Pleistophora vermiformis, Glugea sp., Steinhausia mytilovum and an unidentified microsporidian from E. sinensis, UK. Based on tinctorial properties and background staining, the Luna stain performed better for detection of 6 of the 8 microsporidia. Gram stain was superior for the two microsporidia from invertebrates, Steinhausia mytilovum and the unidentified microsporidian from E. sinensis. PMID:21848126

A vocabulary for the identification and delineation of teratoma tissue components in hematoxylin and eosin-stained samples

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Ramamurthy Bhagavatula

2014-01-01

Full Text Available We propose a methodology for the design of features mimicking the visual cues used by pathologists when identifying tissues in hematoxylin and eosin (H&E-stained samples. Background: H&E staining is the gold standard in clinical histology; it is cheap and universally used, producing a vast number of histopathological samples. While pathologists accurately and consistently identify tissues and their pathologies, it is a time-consuming and expensive task, establishing the need for automated algorithms for improved throughput and robustness. Methods: We use an iterative feedback process to design a histopathology vocabulary (HV, a concise set of features that mimic the visual cues used by pathologists, e.g. "cytoplasm color" or "nucleus density." These features are based in histology and understood by both pathologists and engineers. We compare our HV to several generic texture-feature sets in a pixel-level classification algorithm. Results: Results on delineating and identifying tissues in teratoma tumor samples validate our expert knowledge-based approach. Conclusions: The HV can be an effective tool for identifying and delineating teratoma components from images of H&E-stained tissue samples.

Immunolocalization of transforming growth factor alpha in normal human tissues

DEFF Research Database (Denmark)

Christensen, M E; Poulsen, Steen Seier

1996-01-01

anchorage-independent growth of normal cells and was, therefore, considered as an "oncogenic" growth factor. Later, its immunohistochemical presence in normal human cells as well as its biological effects in normal human tissues have been demonstrated. The aim of the present investigation was to elucidate...... the distribution of the growth factor in a broad spectrum of normal human tissues. Indirect immunoenzymatic staining methods were used. The polypeptide was detected with a polyclonal as well as a monoclonal antibody. The polyclonal and monoclonal antibodies demonstrated almost identical immunoreactivity. TGF......-alpha was found to be widely distributed in cells of normal human tissues derived from all three germ layers, most often in differentiated cells. In epithelial cells, three different kinds of staining patterns were observed, either diffuse cytoplasmic, cytoplasmic in the basal parts of the cells, or distinctly...

Immunohistochemical analysis of medullary breast carcinoma autoantigens in different histological types of breast carcinomas

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Kostianets Olga

2012-11-01

Full Text Available Abstract Background On the past decade a plethora of investigations were directed on identification of molecules involved in breast tumorogenesis, which could represent a powerful tool for monitoring, diagnostics and treatment of this disease. In current study we analyzed six previously identified medullary breast carcinoma autoantigens including LGALS3BP, RAD50, FAM50A, RBPJ, PABPC4, LRRFIP1 with cancer restricted serological profile in different histological types of breast cancer. Methods Semi-quantitative immunohistochemical analysis of 20 tissue samples including medullary breast carcinoma, invasive ductal carcinoma, invasive lobular carcinoma and non-cancerous tissues obtained from patients with fibrocystic disease (each of five was performed using specifically generated polyclonal antibodies. Differences in expression patterns were evaluated considering percent of positively stained cells, insensitivity of staining and subcellular localization in cells of all tissue samples. Results All 6 antigens predominantly expressed in the most cells of all histological types of breast tumors and non-cancerous tissues with slight differences in intensity of staining and subcellular localization. The most significant differences in expression pattern were revealed for RAD50 and LGALS3BP in different histological types of breast cancer and for PABPC4 and FAM50A antigens in immune cells infiltrating breast tumors. Conclusions This pilot study made possible to select 4 antigens LGALS3BP, RAD50, PABPC4, and FAM50A as promising candidates for more comprehensive research as potential molecular markers for breast cancer diagnostics and therapy. Virtual slides The virtual slides’ for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1860649350796892

Oral melanocytic nevi: Report of two cases with immunohistochemical elaboration of their probable origin and maturation

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Dipti Dutta

2015-01-01

Full Text Available Oral melanocytic nevi are localized developmental tissue malformations of nevus cells in the oral mucosa. Relatively rare in occurrence compared to their dermal counterparts, considerable debate exists in the literature related to their origin, development and maturation, and their relationship to oral melanocytes.We report two cases of oral melanocytic nevi with classical clinical presentation. The histopathology was consistent with the known patterns of oral melanocytic nevi. Special stains such as Masson Fontana, further substantiated the observation. S-100 and HMB-45 were applied to immunohistochemically elaborate the cell population. Interestingly two distinct cell populations were detected in the lesions. "Type A" cells in the center of the lesion were S-100 positive, indicating a neural origin and immaturity in development, while peripheral "type B" cells stained positive with HMB-45, indicating melanocytic origin and mature development.

Short Communication: Immunohistochemical localization of the immune cell marker CD68 in bovine adipose tissue: impact of tissue alterations and excessive fat accumulation in dairy cows.

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Häussler, S; Germeroth, D; Laubenthal, L; Ruda, L F; Rehage, J; Dänicke, S; Sauerwein, H

2017-01-01

With the onset of lactation energy from feed intake is mostly insufficient to meet the requirements of dairy cows. Lipid mobilization from adipose tissue (AT) could lead to a compromised inflammatory response enhancing the incidence for diseases. In addition, tissue alterations can occur, displaying areas of necrosis and inflammation. Furthermore, over-conditioned cows mobilizing more lipids from AT than thin cows are prone to develop metabolic disorders. This might lead to an increased infiltration of phagocytic immune cells into AT. In the present study, CD68 positive cells were localized in AT from 10 early lactating Holstein cows displaying different grades of AT alterations. Biopsies were sampled from visceral and subcutaneous AT and the number of CD68 positive cells was immunohistochemically determined. In addition, AT biopsies from over-conditioned, non-pregnant, non-lactating cows (n=8) were immunohistochemically analyzed for CD68 positive cells. The percentage of CD68 positive cells was less than 2% in AT biopsies with tissue alterations and in AT from over-conditioned cows. Therefore, immune cell infiltration demonstrated via the localization of CD68 positive cells seems to play only a minor role in AT from over-conditioned cows as well as in different bovine AT depots with tissue alterations. Copyright © 2016 Elsevier B.V. All rights reserved.

Immunohistochemical Markers Distinguishing Cholangiocellular Carcinoma (CCC) from Pancreatic Ductal Adenocarcinoma (PDAC) Discovered by Proteomic Analysis of Microdissected Cells.

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Padden, Juliet; Ahrens, Maike; Kälsch, Julia; Bertram, Stefanie; Megger, Dominik A; Bracht, Thilo; Eisenacher, Martin; Kocabayoglu, Peri; Meyer, Helmut E; Sipos, Bence; Baba, Hideo A; Sitek, Barbara

2016-03-01

Cholangiocellular carcinoma (CCC) and pancreatic ductal adenocarcinoma (PDAC) are two highly aggressive cancer types that arise from epithelial cells of the pancreatobiliary system. Owing to their histological and morphological similarity, differential diagnosis between CCC and metastasis of PDAC located in the liver frequently proves an unsolvable issue for pathologists. The detection of biomarkers with high specificity and sensitivity for the differentiation of these tumor types would therefore be a valuable tool. Here, we address this problem by comparing microdissected CCC and PDAC tumor cells from nine and eleven cancer patients, respectively, in a label-free proteomics approach. The novel biomarker candidates were subsequently verified by immunohistochemical staining of 73 CCC, 78 primary, and 18 metastatic PDAC tissue sections. In the proteome analysis, we found 180 proteins with a significantly differential expression between CCC and PDAC cells (p value 2). Nine candidate proteins were chosen for an immunohistochemical verification out of which three showed very promising results. These were the annexins ANXA1, ANXA10, and ANXA13. For the correct classification of PDAC, ANXA1 showed a sensitivity of 84% and a specificity of 85% and ANXA10 a sensitivity of 90% at a specificity of 66%. ANXA13 was higher abundant in CCC. It presented a sensitivity of 84% at a specificity of 55%. In metastatic PDAC tissue ANXA1 and ANXA10 showed similar staining behavior as in the primary PDAC tumors (13/18 and 17/18 positive, respectively). ANXA13, however, presented positive staining in eight out of eighteen secondary PDAC tumors and was therefore not suitable for the differentiation of these from CCC. We conclude that ANXA1 and ANXA10 are promising biomarker candidates with high diagnostic values for the differential diagnosis of intrahepatic CCC and metastatic liver tumors deriving from PDAC. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Diagnostic utility of melanin production by fungi: Study on tissue sections and culture smears with Masson-Fontana stain

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Challa Sundaram

2014-01-01

Full Text Available Background: Dematiaceous fungi appear brown in tissue section due to melanin in their cell walls. When the brown color is not seen on routine H and E and culture is not available, differentiation of dematiaceous fungi from other fungi is difficult on morphology alone. Aims and Objective: To study if melanin production by dematiaceous fungi can help differentiate them from other types of fungi. Materials and Methods: Fifty tissue sections of various fungal infections and 13 smears from cultures of different species of fungi were stained with Masson Fontana stain to assess melanin production. The tissue sections included biopsies from 26 culture-proven fungi and 24 biopsies of filamentous fungi diagnosed on morphology alone with no culture confirmation. Results: All culture-proven dematiaceous fungi and Zygomycetes showed strong positivity in sections and culture smears. Aspergillus sp showed variable positivity and intensity. Cryptococcus neoformans showed strong positivity in tissue sections and culture smears. Tissue sections of septate filamentous fungi (9/15, Zygomycetes (4/5, and fungi with both hyphal and yeast morphology (4/4 showed positivity for melanin. The septate filamentous fungi negative for melanin were from biopsy samples of fungal sinusitis including both allergic and invasive fungal sinusitis and colonizing fungal balls. Conclusion: Melanin is produced by both dematiaceous and non-dematiaceous fungi. Masson-Fontana stain cannot reliably differentiate dematiaceous fungi from other filamentous fungi like Aspergillus sp; however, absence of melanin in the hyphae may be used to rule out dematiaceous fungi from other filamentous fungi. In the differential diagnosis of yeast fungi, Cryptococcus sp can be differentiated from Candida sp by Masson-Fontana stain in tissue sections.

Technique and Feasibility of a Dual Staining Method for Estrogen Receptors and AgNORs

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Lukas Günther

2000-01-01

Full Text Available A new staining method for dual demonstration of Estrogen receptors (ER and argyrophilc Nucleolus‐Organizer Regions (AgNORs was developed. To rule out possible reciprocal effects, serial slides of 10 invasive ductale breast cancers were stained with either the single staining method or the simultaneous ER/AgNOR‐staining method and investigated comparatively. By measuring the slides with the image analysis system AMBA, reciprocal effects could be excluded. It was proven that dual staining of both markers results in a reproducible and specific staining result. We concluded that it is justified to measure AgNORs in immunohistochemically stained cells.

Immunohistochemical study of p53 overexpression in radiation-induced colon cancers

International Nuclear Information System (INIS)

Minami, Kazunori; Hayashi, Nobuyuki; Mokarim, A.; Matsuzaki, Sumihiro; Ito, Masahiro; Sekine, Ichiro.

1998-01-01

The expressions of p53 and proliferating cell nuclear antigen (PCNA) were studied immunohistochemically from paraffin sections of 7 cases (9 lesions) of radiation-induced colon cancer and 42 cases of spontaneous colon cancer. Age distribution of radiation-induced and spontaneous colon cancer were 68.1 years (range, 56 to 77 years) and 67.4 years (range, 31 to 85 years), respectively. Among the radiation-induced colon cancers, there were 3 lesions of mucinous carcinoma (33%), a much higher than found for spontaneous mucinous cancer. Immunohistochemically, p53 protein expression was detected in 7/9 (78%) of radiation-induced cancers and in 23/42 (55%) of spontaneous colon cancers. χ 2 analysis found no significant differences between radiation-induced and spontaneous colon cancers in age distribution or p53-positive staining for frequency, histopathology, or Dukes'' classification. In radiation colitis around the cancers including aberrant crypts, spotted p53 staining and abnormal and scattered PCNA-positive staining were observed. In histologically normal cells, p53 staining was almost absent and PCNA-positive staining was regularly observed in the lower half of the crypt. In radiation colitis including aberrant glands, cellular proliferation increased and spotted p53 expression was observed. This study suggests that radiation colitis and aberrant glands might possess malignant potential and deeply associate with carcinogenesis of radiation-induced colon cancer. (author)

Joint Variable Selection and Classification with Immunohistochemical Data

Directory of Open Access Journals (Sweden)

Debashis Ghosh

2009-01-01

Full Text Available To determine if candidate cancer biomarkers have utility in a clinical setting, validation using immunohistochemical methods is typically done. Most analyses of such data have not incorporated the multivariate nature of the staining profiles. In this article, we consider modelling such data using recently developed ideas from the machine learning community. In particular, we consider the joint goals of feature selection and classification. We develop estimation procedures for the analysis of immunohistochemical profiles using the least absolute selection and shrinkage operator. These lead to novel and flexible models and algorithms for the analysis of compositional data. The techniques are illustrated using data from a cancer biomarker study.

Immunohistochemical Characterization of S100A6 in the Murine Ovary

International Nuclear Information System (INIS)

Hanaue, Mayu; Miwa, Naofumi; Takamatsu, Ken

2012-01-01

S100 proteins comprise a large family of Ca 2+ -binding proteins and exhibit a variety of intra- and extracellular functions. Despite our growing knowledge about the biology of S100 proteins in some tissues such as brain and smooth muscle, little is known about S100 proteins in the normal mammalian reproductive tissue. In the present study, we investigated the distribution pattern of S100A6 (alternatively named calcyclin) in the murine ovary by immunohistochemical study using specific antibody. S100A6 was localized substantially in the cytoplasm of luteal cells, with concomitant expression of S100A11, another S100 protein, but not in the other type of cells such as oocytes, follicle epithelial cells (granulosa cells), and cells of stroma including theca interna cells in the murine ovary. S100A6-immunoreactive corpora lutea (CLs) were divided into two types: homogeneously and heterogeneously stained CLs, and possibly they may represent differentiating and mature CL, respectively. Our regression analysis revealed that expression level of S100A6 positively correlated with that of cytochrome P450 11A, a steroidogenic enzyme in the heterogeously stained CL. These results suggested that S100A6 may contribute to differentiation of steroidogenic activity of luteal cells in a synergistic manner with S100A11 by facilitating some shared functions

Computer-assisted assessment of the Human Epidermal Growth Factor Receptor 2 immunohistochemical assay in imaged histologic sections using a membrane isolation algorithm and quantitative analysis of positive controls

International Nuclear Information System (INIS)

Hall, Bonnie H; Ianosi-Irimie, Monica; Javidian, Parisa; Chen, Wenjin; Ganesan, Shridar; Foran, David J

2008-01-01

Breast cancers that overexpress the human epidermal growth factor receptor 2 (HER2) are eligible for effective biologically targeted therapies, such as trastuzumab. However, accurately determining HER2 overexpression, especially in immunohistochemically equivocal cases, remains a challenge. Manual analysis of HER2 expression is dependent on the assessment of membrane staining as well as comparisons with positive controls. In spite of the strides that have been made to standardize the assessment process, intra- and inter-observer discrepancies in scoring is not uncommon. In this manuscript we describe a pathologist assisted, computer-based continuous scoring approach for increasing the precision and reproducibility of assessing imaged breast tissue specimens. Computer-assisted analysis on HER2 IHC is compared with manual scoring and fluorescence in situ hybridization results on a test set of 99 digitally imaged breast cancer cases enriched with equivocally scored (2+) cases. Image features are generated based on the staining profile of the positive control tissue and pixels delineated by a newly developed Membrane Isolation Algorithm. Evaluation of results was performed using Receiver Operator Characteristic (ROC) analysis. A computer-aided diagnostic approach has been developed using a membrane isolation algorithm and quantitative use of positive immunostaining controls. By incorporating internal positive controls into feature analysis a greater Area Under the Curve (AUC) in ROC analysis was achieved than feature analysis without positive controls. Evaluation of HER2 immunostaining that utilized membrane pixels, controls, and percent area stained showed significantly greater AUC than manual scoring, and significantly less false positive rate when used to evaluate immunohistochemically equivocal cases. It has been shown that by incorporating both a membrane isolation algorithm and analysis of known positive controls a computer-assisted diagnostic algorithm was

Pannus tissue at the cartilage-synovium junction in rheumatoid arthritis.

OpenAIRE

Takasugi, Shigeki; Inoue, Hajime

1988-01-01

The cartilage-synovium junction of knees afflicted with rheumatoid arthritis was observed light microscopically using formalin-fixed, decalcified and immunohistochemically stained tissues. Decalcification had little or no influence on immunoreactivity for lysozyme and S-100 protein. All the specimens had pannus formation, which was classified into four types: A) cellular pannus with homogeneous cell pattern, B) cellular pannus of inflammatory cells, C) fibrous pannus with many fibrous bundles...

CYP3A isoforms in Ewing's sarcoma tumours: an immunohistochemical study with clinical correlation.

Science.gov (United States)

Zia, Hamid; Murray, Graeme I; Vyhlidal, Carrie A; Leeder, J Steven; Anwar, Ahmed E; Bui, Marilyn M; Ahmed, Atif A

2015-04-01

Ewing's sarcoma is an aggressive malignancy of bone and soft tissue with high incidence of metastasis and resistance to chemotherapy. Cytochrome P450 (CYP) monooxygenases are a family of enzymes that are involved in the metabolism of exogenous and endogenous compounds, including anti-cancer drugs, and have been implicated in the aggressive behaviour of various malignancies. Tumour samples and clinical information including age, sex, tumour site, tumour size, clinical stage and survival were collected from 36 adult and paediatric patients with Ewing's sarcoma family tumours. Tissue microarrays slides were processed for immunohistochemical labelling for CYP3A4, CYP3A5 and CYP3A7 using liver sections as positive control. The intensity of staining was scored as negative, low or high expression and was analysed statistically for any association with patients' clinical information. Four cases were later excluded due to inadequate viable tissue. CYP3A4 staining was present in 26 (81%) cases with high expression noted in 13 (40%) of 32 cases. High expression was significantly associated with distant metastases (P Ewing's sarcoma tumours and high CYP3A4 expression may be associated with metastasis. Additional studies are needed to further investigate the role of CYP3A4 in the prognosis of these tumours. © 2015 The Authors. International Journal of Experimental Pathology © 2015 International Journal of Experimental Pathology.

The clinicopathologic association of c-MET overexpression in Iranian gastric carcinomas; an immunohistochemical study of tissue microarrays.

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Sotoudeh, Kambiz; Hashemi, Forough; Madjd, Zahra; Sadeghipour, Alireza; Molanaei, Saadat; Kalantary, Elham

2012-05-28

c-MET is an oncogene protein that plays important role in gastric carcinogenesis and has been introduced as a prognostic marker and potential therapeutic target. The aim of this study was to evaluate the frequency of c-MET overexpression and its relationship with clinicopathological variables in gastric cancer of Iranian population using tissue microarray. In a cross sectional study, representative paraffin blocks of 130 patients with gastric carcinoma treated by curative gastrectomy during a 2 years period of 2008-2009 in two university hospitals in Tehran-Iran were collected in tissue microarray and c-MET expression was studied by immunohistochemical staining. Finally 124 cases were evaluated, constituted of 99 male and 25 female with the average age of 61.5 years. In 71% (88/124) of tumors, c-MET high expression was found. c-MET high expression was more associated with intestinal than diffuse tumor type (P = 0.04), deeper tumor invasion, pT3 and pT4 versus pT1 and pT2 (P = 0.014), neural invasion (P = 0.002) and advanced TNM staging, stage 3 and 4 versus stage 1 and2 (P = 0.044). The c-MET high expression was not associated with age, sex, tumor location, differentiation grade and distant metastasis, but relative associations with lymph node metastasis (P = 0.065) and vascular invasion (P = 0.078) were observed. c-MET oncogene protein was frequently overexpressed in Iranian gastric carcinomas and it was related to clinicopathological characteristics such as tumor type, depth of invasion, neural invasion and TNM staging. It can also support the idea that c-MET is a potential marker for target therapy in Iranian gastric cancer. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9744598757151429.

Fat tissue after lipolysis of lipomas: a histopathological and immunohistochemical study.

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Bechara, Falk G; Sand, Michael; Hoffmann, Klaus; Sand, Daniel; Altmeyer, Peter; Stücker, Markus

2007-07-01

Injections with Lipostabil, a phosphatidylcholine (PDC) containing substance, have become a popular technique to treat localized fat accumulation and lipomas for aesthetic reasons. Despite its frequent use, the mechanism of action of PDC and histological changes of treated fat tissue still remain unclear. To investigate the histological changes of lipomas after treatment with PDC. In all, fourteen lipomas (n = 14) in five patients presenting with multiple lipomas were treated with intralesional injections of PDC (Lipostabil, Nettermann, Germany). Histological changes with immunohistochemical analysis of the inflammatory process were evaluated 4, 10, 24, 48 h, 10 days, 30 days and 60 days after lipolysis. Between 4 and 48 h after injection, histology shows a lobular neutrophilic infiltrate with partially destroyed fat cells. At day 10 the inflammatory process is accompanied by an infiltration of T-lymphocytes. After 60 days formation of macrophages with foam cells are visible, accompanied by thickened septa and capsula. Lipolysis with PDC results in a distinct inflammatory reaction of affected fat tissue, similar to factitial panniculitis. Early destruction of fat cells may suggest the involvement of detergent or osmotic mechanisms in the process.

Endometriosis-associated hydrocele of the canal of Nuck with immunohistochemical confirmation: a case report.

Science.gov (United States)

Okoshi, Kae; Mizumoto, Masaki; Kinoshita, Koichi

2017-12-21

The canal of Nuck is an embryological vestige of the processus vaginalis, and presents a potential site for endometriosis seeding. Hydroceles in this region are a rare cause of inguinal swelling in females. In addition, endometriosis localized to the canal of Nuck is exceedingly rare. A 44-year-old Japanese woman presented with a painful mass overlying her right pubis. She underwent surgery to completely excise the mass. During surgery, division of the external oblique aponeurosis revealed a cyst that occupied the inguinal canal and it adhered to the transverse fascia, inguinal ligament, and pubic bone. The cyst was dissected from the round ligament, and the defect in the internal inguinal ring was repaired and reinforced with mesh. On macroscopic examination, the cyst had a heterogeneous fibrous aspect with dark brown inclusions. Microscopic examination revealed that the cyst was tortuous, lined by mesothelial-like cells, and accompanied by partial subcapsular hemorrhage. Endometrium-like tissue was observed in the cystic wall. Immunohistochemical staining for podoplanin confirmed the mesothelial origin of the cyst-lining cells. The epithelial cells and stromal cells were positive for estrogen receptors. In this case of an endometriosis-associated hydrocele of the canal of Nuck, the mesothelial origin of the cyst-lining cells and endometriosis were confirmed by positive immunohistochemical staining for podoplanin and estrogen receptors, respectively. We determined that hydrocele resection and reinforcement of the anterior inguinal canal wall (if necessary) are appropriate treatments for this condition.

Double epi-illumination microscopy with separate visualization of two antigens: a combination of epi-polarization for immunogold-silver staining and epi-fluorescence for alkaline phosphatase staining

NARCIS (Netherlands)

van der Loos, C. M.; Becker, A. E.

1994-01-01

We present a method for an epi-illumination immunohistochemical double staining approach. The method combines the use of an immuno-alkaline phosphatase technique and the immunogold-silver technique, visualized with epifluorescence and epi-polarization illumination, respectively. Out of six tested

Gram staining.

Science.gov (United States)

Coico, Richard

2005-10-01

Named after Hans Christian Gram who developed the method in 1884, the Gram stain allows one to distinguish between Gram-positive and Gram-negative bacteria on the basis of differential staining with a crystal violet-iodine complex and a safranin counterstain. The cell walls of Gram-positive organisms retain this complex after treatment with alcohol and appear purple, whereas gram-negative organisms decolorize following such treatment and appear pink. The method described here is useful for assessing bacterial contamination of tissue culture samples or for examining the Gram stain status and morphological features of bacteria isolated from mixed or isolated bacterial cultures.

Up-regulation of ALG-2 in hepatomas and lung cancer tissue

DEFF Research Database (Denmark)

la Cour, Jonas Marstrand; Mollerup, Jens; Winding, Pernille

2003-01-01

, a result confirmed by immunohistochemical analysis. Staining of four different lung cancer tissue microarrays including specimens of 263 patients showed that ALG-2 is mainly localized to epithelial cells and significantly up-regulated in small-cell lung cancers and in non-small-cell lung cancers. Our...... using Western blot analysis and immunohistochemistry. Western blot analysis of 15 different adult mouse tissues demonstrated that ALG-2 is ubiquitously expressed. We found that ALG-2 was more than threefold overexpressed in rat liver hepatoma compared to normal rat liver using Western blot analysis...

Immunohistochemically determined total epidermal growth factor receptor levels not of prognostic value in newly diagnosed glioblastoma multiforme: Report from the Radiation Therapy Oncology Group

International Nuclear Information System (INIS)

Chakravarti, Arnab; Seiferheld, Wendy; Tu Xiaoyu; Wang Huijun; Zhang Huazhong; Ang, K. Kian; Hammond, Elizabeth; Curran, Walter; Mehta, Minesh

2005-01-01

Purpose: The Radiation Therapy Oncology Group (RTOG) performed an analysis of patterns of immunohistochemically detected total epidermal growth factor receptor (EGFR) protein expression levels and their prognostic significance on archival tissue in newly diagnosed glioblastoma multiforme (GBM) patients from prior prospective RTOG clinical trials. Methods and materials: Patients in this study had been treated on previous RTOG GBM trials (RTOG 7401, 7918, 8302, 8409, 9006, 9305, 9602, and 9806). Tissue microarrays were prepared from 155 patients enrolled in these trials. These specimens were stained using a mouse monoclonal antibody specific for the extracellular binding domain of EGFR to detect total EGFR (including both wild-type phosphorylated and wild-type unphosphorylated isoforms with some cross-reactivity with EGFRvIII). The intensity of total EGFR protein expression was measured by computerized quantitative image analysis using the SAMBA 4000 Cell Image Analysis System. The parameters measured were the mean optical densities over the labeled areas and the staining index, which represents the proportion of stained area relative to the mean stain concentration. Both parameters were correlated with the clinical outcome. Results: No differences in either overall or progression-free survival could be demonstrated by the mean optical density class or mean optical density quartile or the staining index of total EGFR immunostaining among the representative RTOG GBM cases. Conclusion: Total EGFR protein expression levels, as measured immunohistochemically, do not appear to be of prognostic value in newly diagnosed GBM patients. Given the accumulating clinical evidence of the activity of anti-EGFR agents in GBM and the preclinical data suggesting the important role of downstream mediators as effectors of EGFR signaling, the RTOG is conducting additional investigations into the prognostic value of activation patterns of EGFR signaling, both at the level of the receptor

Pitfalls using tyramide signal amplification (TSA) in the mouse gastrointestinal tract: endogenous streptavidin-binding sites lead to false positive staining.

Science.gov (United States)

Horling, L; Neuhuber, W L; Raab, M

2012-02-15

Highly sensitive immunohistochemical detection systems such as tyramide signal amplification (TSA) are widely used, since they allow using two primary antibodies raised in the same species. Most of them are based on the streptavidin-biotin-peroxidase system and include streptavidin-coupled secondary antibodies. Using TSA in cryostat-sectioned tissues of mouse esophagus, we were puzzled by negative controls with unexpected staining mostly in the ganglionic areas. This prompted us to search for the causing agent and to include also other parts of the mouse gastrointestinal tract for comparison. Streptavidin-coupled antibodies bound to endogenous binding sites yet to be characterized, which are present throughout the mouse intestines. Staining was mainly localized around neuronal cell bodies of enteric ganglia. Thus, caution is warranted when applying streptavidin-coupled antibodies in the mouse gastrointestinal tract. The use of endogenous biotin-blocking kits combined with a prolonged post-fixation time could significantly reduce unintentional staining. Copyright © 2011 Elsevier B.V. All rights reserved.

Histopathologic and immunohistochemical features of Hashimoto thyroiditis.

Science.gov (United States)

Amani, H Kazem

2011-01-01

Intrathyroid lymphoid tissue is accrued in Hashimoto thyroiditis (HT). Histologically, this acquired lymphoid tissue bears a close resemblance to mucosa-associated lymphoid tissue (MALT) and can evolve to lymphoma. To demonstrate the morphological, and immunohistochemical profiles of Hashimoto thyroiditis and to ascertain the importance of light chain restriction in distinguishing HT with extensive lymphoplasmacytoid infiltrate from MALT lymphoma. We studied histopathologically and immunohistochemically (CD20, CD3, Igk, Igl and cytokeratin) 30 cases of HT for evaluation of the lymphoid infiltrate and the presence of lymphoepithelial lesions (LELs). Distinguishing between early thyroid lymphoma and HT was evaluated by light chain restriction. These findings were compared with two cases of primary thyroid lymphoma. The histopathological findings were characteristic of HT. Immunohistochemistry confirmed inconspicuous, rare B-cell LELs as well as a prominent T-lymphocyte population. Testing for light chain restriction showed polyclonal population of plasma cells. The cases of MALT lymphoma had distinct destructive lymphoepithelial lesions, B-cell immunophenotyping and showed kappa light chain restriction in the plasmacytoid population. Hashimoto thyroiditis differs both histopathologically and immunohistochemically from thyroid lymphoma. In suspicious cases, immunohistochemistry could be helpful in reaching a definitive diagnosis.

Expression analysis of polyphenol oxidase isozymes by active staining method and tissue browning of head lettuce (Lactuca sativa L.).

Science.gov (United States)

Noda, Takahiro; Iimure, Kazuhiko; Okamoto, Shunsuke; Saito, Akira

2017-08-01

Browning of plant tissue is generally considered attributable to enzymatic oxidation by polyphenol oxidase (PPO). Electrophoresis followed by activity staining has been used as an effective procedure to visually detect and isolate isozymes; however, it has not been applied for examination of various PPO isozymes in lettuce. Our study demonstrated that different lettuce PPO isozymes could be detected at different pH in active staining, and multiple isozymes were detected only under alkaline conditions. As a result, we concluded that activity staining with approximately pH 8 enabled to detect various PPO isozymes in lettuce. By expression analysis of the PPO isozymes after wounding, PPO isozymes that correlated with time-course of tissue browning were detected. The wound-induced PPO may play a key role in enzymatic browning.

Identification of immune cell infiltration in hematoxylin-eosin stained breast cancer samples: texture-based classification of tissue morphologies

Science.gov (United States)

Turkki, Riku; Linder, Nina; Kovanen, Panu E.; Pellinen, Teijo; Lundin, Johan

2016-03-01

The characteristics of immune cells in the tumor microenvironment of breast cancer capture clinically important information. Despite the heterogeneity of tumor-infiltrating immune cells, it has been shown that the degree of infiltration assessed by visual evaluation of hematoxylin-eosin (H and E) stained samples has prognostic and possibly predictive value. However, quantification of the infiltration in H and E-stained tissue samples is currently dependent on visual scoring by an expert. Computer vision enables automated characterization of the components of the tumor microenvironment, and texture-based methods have successfully been used to discriminate between different tissue morphologies and cell phenotypes. In this study, we evaluate whether local binary pattern texture features with superpixel segmentation and classification with support vector machine can be utilized to identify immune cell infiltration in H and E-stained breast cancer samples. Guided with the pan-leukocyte CD45 marker, we annotated training and test sets from 20 primary breast cancer samples. In the training set of arbitrary sized image regions (n=1,116) a 3-fold cross-validation resulted in 98% accuracy and an area under the receiver-operating characteristic curve (AUC) of 0.98 to discriminate between immune cell -rich and - poor areas. In the test set (n=204), we achieved an accuracy of 96% and AUC of 0.99 to label cropped tissue regions correctly into immune cell -rich and -poor categories. The obtained results demonstrate strong discrimination between immune cell -rich and -poor tissue morphologies. The proposed method can provide a quantitative measurement of the degree of immune cell infiltration and applied to digitally scanned H and E-stained breast cancer samples for diagnostic purposes.

NM23 protein expression in colorectal carcinoma using TMA (tissue microarray: association with metastases and survival

Directory of Open Access Journals (Sweden)

Levindo Alves de Oliveira

2010-12-01

Full Text Available CONTEXT: NM23, a metastasis suppressor gene, may be associated with prognosis in patients with colorectal carcinoma. OBJECTIVE: To analyze NM23 expression and its association with the presence of lymph node and liver metastases and survival in patients operated on for colorectal carcinoma. METHODS: One hundred thirty patients operated on for colorectal carcinoma were investigated. Tissue microarray blocks containing neoplastic tissue and tumor-adjacent non-neoplastic mucosa were obtained and analyzed by immunohistochemical staining using a monoclonal anti-NM23 antibody. Immunohistochemical expression was assessed using a semiquantitative scoring method, counting the percentage of stained cells. The results were compared regarding morphological and histological characteristics of the colorectal carcinoma, presence of lymph node and liver metastases, tumor staging, and patient survival. Statistical analysis was performed using the Mann-Whitney test, the Kruskal-Wallis test and Fisher's exact test. Survival analysis was performed using the Kaplan-Meier method and the log-rank test. RESULTS: NM23 expression was higher in colorectal carcinoma tissue than in adjacent non-neoplastic mucosa (P<0.0001. NM23 protein expression did not correlate with degree of cell differentiation (P = 0.57, vascular invasion (P = 0.85, lymphatic invasion (P = 0.41, perineural infiltration (P = 0.46, staging (P = 0.19, lymph node metastases (P = 0.08, or liver metastases (P = 0.59. Disease-free survival showed significant association (P = 0.01 with the intensity of NM23 protein immunohistochemical expression in colorectal carcinoma tissue, whereas overall survival showed no association with NM23 protein expression (P = 0.13. CONCLUSIONS: NM23 protein expression was higher in neoplastic colorectal carcinoma tissue than in adjacent non-neoplastic mucosa, showing no correlation with morphological aspects, presence of lymph node or liver metastases, colorectal carcinoma

Novel methods of cytokine detection: Real-time PCR, ELISPOT, and intracellular cytokine staining

Directory of Open Access Journals (Sweden)

Eliza Turlej

2009-05-01

Full Text Available Cytokines are small hormone-like proteins that play important roles in immune system control. Cytokines regulate the proliferation and differentiation of cells and hematopoiesis and act as mediators in the inflammatory reaction. Changes in cytokine levels are found in many diseases, such as sepsis, bowel inflammatory disease, autoimmune diseases, as well as graft-versus-host disease. Cytokines levels can be detected using in vivo, in vitro, and ex vivo techniques. The level of cytokine produced can be measured by immunoenzymatic test (ELISA in supernatant after cell culture with the addition of stimulant and in plasma by techniques that measure the level of cytokine secretion in cells (e.g. immunohistochemical staining, ELISPOT, and intracellular cytokine staining, and by molecular biological methods (RPA, real-time PCR, in situ hybridization, and Northern blot. Detection of cytokine mRNA in tissues is useful in the direct determination of heterogenic populations of cytokine-producing cells. Nowadays the most frequently used methods for measuring cytokine level are ELISPOT, intracellular cytokine staining with flow cytometry detection, and real-time PCR. These methods have an important clinical role in vaccine efficacy, in viral, bacterial, and verminous diagnostics, and in determining the efficacy of cancer treatment.

Multispectral Enhancement Method to Increase the Visual Differences of Tissue Structures in Stained Histopathology Images

Directory of Open Access Journals (Sweden)

Pinky A. Bautista

2012-01-01

Full Text Available In this paper we proposed a multispectral enhancement scheme in which the spectral colors of the stained tissue-structure of interest and its background can be independently modified by the user to further improve their visualization and color discrimination. The colors of the background objects are modified by transforming their N-band spectra through an NxN transformation matrix, which is derived by mapping the representative samples of their original spectra to the spectra of their target colors using least mean square method. On the other hand, the color of the tissue structure of interest is modified by modulating the transformed spectra with the sum of the pixel’s spectral residual-errors at specific bands weighted through an NxN weighting matrix; the spectral error is derived by taking the difference between the pixel’s original spectrum and its reconstructed spectrum using the first M dominant principal component vectors in principal component analysis. Promising results were obtained on the visualization of the collagen fiber and the non-collagen tissue structures, e.g., nuclei, cytoplasm and red blood cells (RBC, in a hematoxylin and eosin (H&E stained image.

Naturally acquired bovine besnoitiosis: histological and immunohistochemical findings in acute, subacute, and chronic disease.

Science.gov (United States)

Langenmayer, M C; Gollnick, N S; Majzoub-Altweck, M; Scharr, J C; Schares, G; Hermanns, W

2015-05-01

The pathogenesis of bovine besnoitiosis, a disease of increasing concern within Europe, is still incompletely understood. In this study, disease progression after natural infection with the causative apicomplexan Besnoitia besnoiti was monitored in histological skin sections of 5 individual female cattle over time. High-frequency skin sampling of 2 cattle with mild and 2 with severe acute, subacute, and chronic besnoitiosis, as well as from 1 animal during subclinical disease, enabled documentation from the beginning of the disease. Skin sections were stained with hematoxylin and eosin and Giemsa, periodic acid-Schiff reaction, and anti-Besnoitia immunohistochemistry. In all 4 clinically affected animals, tachyzoite-like endozoites could be detected for the first time by immunohistochemistry, and tissue cyst evolution was monitored. Besnoitiosis-associated lesions were not detected in the animal showing the subclinical course. Because of the inconsistency of the nomenclature of Besnoitia tissue cyst layers in the literature, a new nomenclature for B. besnoiti cyst wall layers is proposed: tissue cysts consist of a hypertrophied host cell with enlarged nuclei, an intracytoplasmic parasitophorous vacuole with bradyzoites, a sometimes vacuolated inner cyst wall, and an outer cyst wall in more developed cysts. Inner and outer cyst walls can be readily distinguished by using special stains. In 1 animal, extracystic B. besnoiti zoites were immunohistochemically detected during the chronic stage. At necropsy, the 2 severely affected cows displayed large numbers of B. besnoiti cysts in a variety of tissues, including the corium of the claws, contributing mainly to the development of chronic laminitis in these 2 cases. © The Author(s) 2014.

Cooperation of Indian Hedgehog and Vascular Endothelial Growth Factor in Tumor Angiogenesis and Growth in Human Hepatocellular Carcinomas, an Immunohistochemical Study.

Science.gov (United States)

Li, Yang; Liu, Yang; Wang, Guangxi; Wang, Yuxiang; Guo, Limei

2018-04-07

The Hedgehog pathway was recently shown to be involved in vascular development and neovascularization in human embryogenesis and disease. However, the role of Hedgehog pathway in modulating tumor angiogenesis is still unexplored. In the current study, we investigated the expression of Indian Hedgehog (Ihh) and vascular endothelial cell growth factor (VEGF) in human hepatocellular carcinomas (HCCs) with immunohistochemical staining and compared the immunoreaction data with various clinicopathologic characteristics. Immunoreactivity of Ihh and VEGF proteins was observed in 61.5% (56/91) and 64.5% (59/91) cases of HCC tumor tissues, respectively, which was considerably higher than the adjacent nonmalignant tissues. Ihh protein was observed predominantly in the cytoplasm of the tumor cells with a staining pattern of which was sparse and dot-like, or circular around the cell membrane. VEGF protein was expressed heterogenously in the cytoplasm in tumor cells and was negative in peritumoral areas in all cases. CD34 showed diffuse staining in the tumor parenchyma in most HCC specimens. The association of expression of Ihh and VEGF with tumor size was statistically significant (PIhh and VEGF proteins in HCC (r=0.6, PIhh and CD34 staining (r=0.261, P=0.012). Our findings suggest that Ihh is involved in the development of HCC. These findings are also consistent with the concept that cooperation of Ihh and VEGF modulate HCC tumor angiogenesis and growth.

Validation of 31 of the most commonly used immunohistochemical antibodies in cytology prepared using the Cellient(®) automated cell block system.

Science.gov (United States)

Montgomery, Eric; Gao, Chen; de Luca, Julie; Bower, Jessie; Attwood, Kristropher; Ylagan, Lourdes

2014-12-01

The Cellient(®) cell block system has become available as an alternative, partially automated method to create cell blocks in cytology. We sought to show a validation method for immunohistochemical (IHC) staining on the Cellient cell block system (CCB) in comparison with the formalin fixed paraffin embedded traditional cell block (TCB). Immunohistochemical staining was performed using 31 antibodies on 38 patient samples for a total of 326 slides. Split samples were processed using both methods by following the Cellient(®) manufacturer's recommendations for the Cellient cell block (CCB) and the Histogel method for preparing the traditional cell block (TCB). Interpretation was performed by three pathologists and two cytotechnologists. Immunohistochemical stains were scored as: 0/1+ (negative) and 2/3+ (positive). Inter-rater agreement for each antibody was evaluated for CCB and TCB, as well as the intra-rater agreement between TCB and CCB between observers. Interobserver staining concordance for the TCB was obtained with statistical significance (P Cellient system are reliable and concordant with stains performed on the same split samples processed via a formalin fixed-paraffin embedded (FFPE) block. The Cellient system is a welcome adjunct to cytology work-flow by producing cell block material of sufficient quality to allow the use of routine IHC. © 2014 Wiley Periodicals, Inc.

Is hematoxylin-eosin staining in rectal mucosal and submucosal biopsies still useful for the diagnosis of Hirschsprung disease?

Science.gov (United States)

Serafini, Suellen; Santos, Maria Mercês; Aoun Tannuri, Ana Cristina; Zerbini, Maria Claudia Nogueira; de Mendonça Coelho, Maria Cecília; de Oliveira Gonçalves, Josiane; Tannuri, Uenis

2017-12-06

Hematoxylin-eosin (HE) staining of a full-thickness rectal wall fragment is classically used for the diagnosis of Hirschsprung disease (HD). However, this technique requires large fragments for a better diagnosis. Additionally, the histochemical and immunohistochemical methods of staining small fragments of rectal mucosal and submucosal biopsies are not available in all centers. Therefore, the possibility of diagnosing HD through HE staining in these biopsies could be a valuable alternative for centers that do not have more specific techniques. The objectives of the current investigation were to evaluate the concordance of the results obtained by HE staining and the calretinin method with acetylcholinesterase (AChE) activity in fragments of mucosa and submucosa in the diagnosis of HD. For this study, 50 cases from our laboratory were selected. The tissue material was embedded in paraffin. Sixty levels of each fragment were utilized for HE, and the other 3 levels were used for calretinin. These slides were analyzed under the microscope, photographed and classified as either positive for HD when no ganglion cells were found with nerve trunks present or as negative when ganglion cells were found. The results from reading the slides were compared with those of AChE. Of the 50 cases evaluated by the HE technique, only 5 contradicted the diagnosis based on AChE, with a Kappa value of 0.800 and an accuracy of 90%. In the comparison between calretinin and AChE, 8 cases were discordant, with a Kappa value of 0.676 and an accuracy of 84%. The concordance of results from AChE and HE methods was satisfactory, allowing for the potential use of the HE method for fragments of mucosa and submucosa as a valid alternative in the diagnosis of HD. The immunohistochemical technique of calretinin did not show good agreement with the AChE activity in our study.

Fluorescence microscopy for the evaluation of elastic tissue patterns within fibrous proliferations of the skin on hematoxylin-eosin-stained slides.

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Borucki, Robert; Perry, David M; Lopez-Garcia, Dan R; Kazlouskaya, Viktoryia; Elston, Dirk M

2018-01-05

Diagnosis of fibrous tumors can be challenging and expensive due to the use of special stains. Determine the usefulness of fluorescence microscopy in the evaluation of elastic pattern on H&E stained slides. A total of 228 slides were evaluated by fluorescence microscopy for elastic tissue patterns and sensitivity and specificity determined for relevant comparisons. Fluorescence microscopy was found to be useful especially in the case of distinguishing dermatofibroma (DF) vs dermatofibrosarcoma protuberans (DFSP) and also dermatomyofibroma (DMF) vs other fibrous tumors. In some cases, excessive background staining made it difficult to interpret. Evaluation of elastic tissue patterns by fluorescence microscopy in fibrous tumors is a cheap and efficient means to further delineate these often challenging tumors. Copyright © 2018. Published by Elsevier Inc.

Hidradenocarcinoma: a histological and immunohistochemical study.

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Ko, Christine J; Cochran, Alistair J; Eng, William; Binder, Scott W

2006-11-01

The diagnosis of hidradenocarcinoma is difficult due to a combination of factors including inconsistent nomenclature/ classification, rarity of the neoplasm, and variable morphology of cells composing the neoplasm. Immunohistochemistry has not been previously performed on a series of hidradenocarcinomas. We evaluated six cases of hidradenocarcinoma histologically and immunohistochemically using antibodies to gross cystic disease fluid protein-15 (GCDFP-15), carcino-embryonic antigen (CEA), epithelial membrane antigen (EMA), S-100 protein, keratin AE1/3, cytokeratin 5/6, p53, bcl-1, bcl-2, and Ki67. Histology suggested concurrent eccrine and apocrine differentiation of the cases. Ki67 and p53 staining was strongly positive in five of six tumors. The neoplasms stained with antibodies to CEA, S-100 protein, GCDFP-15, EMA, bcl-1, and bcl-2 in no consistent pattern. All tumors studied stained positively for keratin AE1/3 and cytokeratin 5/6. In making the diagnosis of hidradenocarcinoma, it may be unnecessary to separate hidradenocarcinoma into eccrine and apocrine categories, and although Ki67 and p53 may be helpful, histological parameters remain paramount.

Immunohistochemical expression of EGFR in colorectal carcinoma correlates with high but not low level gene amplification, as demonstrated by CISH.

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Hemmings, Chris; Broomfield, Amy; Bean, Elaine; Whitehead, Martin; Yip, Desmond

2009-01-01

To assess and compare immunohistochemical expression of epidermal growth factor receptor (EGFR) with gene amplification as demonstrated by chromogenic in situ hybridisation (CISH), in colorectal adenocarcinoma. Sections from 100 consecutive colorectal cancer resection specimens were stained for EGFR using immunohistochemistry and CISH. Immunohistochemical assessment was independently performed at two laboratories, using the same antibody and protocols. With immunohistochemistry, strong circumferential membrane staining (3+ staining) was demonstrated in only 5% of cases, and this was only focal in three of five cases. At one laboratory, weak or incomplete staining (1+ or 2+) was observed in five further cases (5%), which had been negative at the other laboratory. CISH demonstrated high level gene amplification (>10 copies/nucleus) in the same five cases which had demonstrated 3+ staining with immunohistochemistry, and in those cases where the staining was focal, the amplification was demonstrated in the same foci of the tumour. Five further cases (5%) had low level amplification (5-10 copies per nucleus); these cases did not exhibit significant positive staining with immunohistochemistry. All the cases which demonstrated gene amplification (high or low level) arose in the distal colon. There was no correlation between gene amplification status and a variety of other variables, including stage at diagnosis, mucinous differentiation, neuroendocrine differentiation, or loss of expression of mismatch repair proteins. Immunohistochemical expression of EGFR is variable between laboratories, even using standardised protocols. 3+ staining is predictive of high level gene amplification, but correlates very poorly with low level amplification, which may still be clinically significant. In some cases gene amplification was only focal, offering a potential explanation for poor response to targeted therapy in patients with EGFR positive tumours.

Immunohistochemical Characterization of Canine Lymphomas

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Roxana CORA

2017-11-01

Full Text Available Lymphomas occur by clonal expansion of lymphoid cells and have distinctive morphological and immunophenotypic features. Determination of canine lymphoma immunophenotype is useful for accurate prognosis and further therapy. In the suggested study, we performed an immunohistochemical evaluation of some cases with canine lymphoma diagnosed in the Department of Pathology (Faculty of Veterinary Medicine, Cluj-Napoca, Romania, in order to characterize them. The investigation included 39 dogs diagnosed with different anatomical forms of lymphoma, following necropsy analysis or assessment of biopsies. The diagnosis of lymphoma was confirmed by necropsy and histopathology (Hematoxylin-eosin stain examinations. The collected specimens were analyzed by immunohistochemistry technique (automatic method using the following antibodies: CD3, CD20, CD21 and CD79a. The analyzed neoplasms were characterized as follows: about 64.10% of cases were diagnosed as B-cell lymphomas, 33.34% of cases as T-cell lymphomas, whereas 2.56% of cases were null cell type lymphomas (neither B nor T. Most of multicentric (80%, mediastinal (60% and primary central nervous system lymphomas (100% had B immunophenotype, while the majority of cutaneous (80% and digestive (100% lymphomas had T immunophenotype. Immunohistochemical description of canine lymphomas can deliver some major details concerning their behavior and malignancy. Additionally, vital prognosis and efficacy of some therapeutic protocols are relying on the immunohistochemical features of canine lymphoma.

[Histochemical stains for minerals by hematoxylin-lake method].

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Miyagawa, Makoto

2013-04-01

The present study was undertaken to establish the experimental animal model by histological staining methods for minerals. After intraperitoneal injections of minerals, precipitates deposited on the surface of the liver. Liver tissues were fixed in paraformaldehyde, embedded in paraffin and cut into thin sections which were used as minerals containing standard section. Several reagents for histological stains and spectrophotometry for minerals were applied in both test-tube experiments and stainings of tissue sections to test for minerals. Hematoxylin-lake was found of capable of staining minerals in tissue. A simple technique used was described for light microscopic detection of minerals.

Hidradenocarcinoma presenting as soft tissue mass: Case report with cytomorphologic description, histologic correlation, and differential diagnosis.

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Jinnah, Alexander H; Emory, Cynthia L; Mai, Nicholas H; Bergman, Simon; Salih, Ziyan T

2016-05-01

Hidradenocarcinoma (HAC) is a rare adenexal tumor with a propensity for the head and neck region and extremities. We report a case of hidradenocarcinnoma in a 56-year-old woman with a mass on her right palm sampled by fine-needle aspiration and later confirmed on histological examination. Fine-needle aspiration cytology revealed a dual population of cells including polyhedral eosinophilic cells and glycogen containing cells with pale/clear cytoplasm. The nuclei were pleomorphic with prominent nucleoli. Occassional papillary structures were identified on the cell block material. A series of immunohistochemical stains were performed and an adnexal neoplasm was suggested. The mass was resected. On histologic sections, infiltration into the adjacent soft tissue was identified. After an additional series of immunohistochemical stains, the diagnosis was confirmed as a HAC. Herein, we present our findings and discuss the differential diagnoses. © 2016 Wiley Periodicals, Inc.

Computer-assisted assessment of the Human Epidermal Growth Factor Receptor 2 immunohistochemical assay in imaged histologic sections using a membrane isolation algorithm and quantitative analysis of positive controls

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Ianosi-Irimie Monica

2008-06-01

Full Text Available Abstract Background Breast cancers that overexpress the human epidermal growth factor receptor 2 (HER2 are eligible for effective biologically targeted therapies, such as trastuzumab. However, accurately determining HER2 overexpression, especially in immunohistochemically equivocal cases, remains a challenge. Manual analysis of HER2 expression is dependent on the assessment of membrane staining as well as comparisons with positive controls. In spite of the strides that have been made to standardize the assessment process, intra- and inter-observer discrepancies in scoring is not uncommon. In this manuscript we describe a pathologist assisted, computer-based continuous scoring approach for increasing the precision and reproducibility of assessing imaged breast tissue specimens. Methods Computer-assisted analysis on HER2 IHC is compared with manual scoring and fluorescence in situ hybridization results on a test set of 99 digitally imaged breast cancer cases enriched with equivocally scored (2+ cases. Image features are generated based on the staining profile of the positive control tissue and pixels delineated by a newly developed Membrane Isolation Algorithm. Evaluation of results was performed using Receiver Operator Characteristic (ROC analysis. Results A computer-aided diagnostic approach has been developed using a membrane isolation algorithm and quantitative use of positive immunostaining controls. By incorporating internal positive controls into feature analysis a greater Area Under the Curve (AUC in ROC analysis was achieved than feature analysis without positive controls. Evaluation of HER2 immunostaining that utilized membrane pixels, controls, and percent area stained showed significantly greater AUC than manual scoring, and significantly less false positive rate when used to evaluate immunohistochemically equivocal cases. Conclusion It has been shown that by incorporating both a membrane isolation algorithm and analysis of known

Hemangiopericytoma in a young dog: Evaluation of histopathological and immunohistochemical features

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Fatemeh Namazi

2014-06-01

Full Text Available In the present study, we describe a subcutaneous mass between the left flank and hip in a 2-year-old male Great Dane dog. Histopathologically, cells appeared to be spindle shaped around a central capillary together with a fingerprint pattern. Immunohistochemical analysis presented that the neoplastic cells expressed vimentin, but did not stain for S-100 protein. On the basis of histopathology and immunohistochemical findings, the present tumor was diagnosed as canine hemangiopericytoma. Hemangiopericytoma could be considered in differential diagnosis list of any mass in the skin (even in young dogs and must be identified histopathologically.

Lipid-rich histology in a basal-type immunoprofile breast carcinoma: histochemical and immunohistochemical analysis of a case

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Francesco Maria Maiello

2009-12-01

Full Text Available We describe the clinicopathological and morphological features of an unusual breast carcinoma classifiable as a lipid-rich variant of ductal invasive carcinoma, with a basal-type immunohistochemical profile. Basal-type breast cancers show no hormonal receptor expression, rarely over-express HER-2 but exhibit molecular high weight cytokeratins, EGFR and c-kit positivity. Special stains and histochemistry tests were used to elucidate the nature of vescicles in the neoplastic cells. Sudan IV was performed on formalin-fixed tissue. Commercially available antibodies tested were: ER, PgR, EGFR, HER2, c-kit, high molecular weight cytokeratins. Cytoplasmic lipids were highlighted as red-orange droplets on Sudan IV staining. As for immunohistochemistry, the tumor showed no reactivity to ER, PgR and HER2 (triple negative, and diffuse and strong positivity to high weight cytokeratins, EGFR and c-kit, such as a basal-type breast carcinoma. A basaloid phenotype in a lipid-rich carcinoma has not been previously reported.

Histomorphological and immunohistochemical characterization of 172 cutaneous round cell tumours in dogs

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Marina Rios Araújo

2012-08-01

Full Text Available This paper describes the use of a panel of antibodies (CD117, CD3, CD79a, CD45, cytokeratin, vimentin and E-cadherin on formalin-fixed, paraffin-embedded sections of canine cutaneous round cell tumours. Neoplastic tumours were diagnosed by histology and histochemical stains and included 107 mast cell tumours, 31 cutaneous histiocytomas, two localized histiocytic sarcomas, 21 cutaneous lymphomas, three plasma cell tumours, one transmissible venereal tumour and seven unclassified round cell tumours. The histologic diagnosis was modified in 39.5% of the total 172 neoplasms. The staining for CD45 and Ecadherin were variable, and therefore, the final diagnoses of cutaneous histiocytoma and localized histiocytic sarcoma were made based on histology in association with negative results for CD3, CD79a, CD117 and cytokeratin. The cellular origin of unclassified round cell tumours was defined in all cases. Cutaneous B-cell lymphoma and plasma cell tumours were CD79a-positive and could be distinguished from each other by the morphological characteristics. Mast cell tumours and T cell lymphoma were CD117 and CD3 positive, respectively. The positive staining for vimentin and the negative staining for CD3, CD79a, CD117 and cytokeratin favoured the diagnosis of transmissible venereal tumours. Thus, the final diagnosis of cutaneous round cell tumours should be based on the interpretation of immunohistochemical results together with the cellular morphology observed by histology. Therefore, more studies to optimize the specific markers in formalin-fixed, paraffinembedded tissues (especially for histiocytes are required for definitive diagnosis of round cell tumours in dogs.

Evaluation of Calretinin expression in Ameloblastoma and Non-Neoplastic Odontogenic Cysts - An immunohistochemical study.

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D'Silva, Shaloom; Sumathi, M K; Balaji, N; Shetty, Nisha K N; Pramod, K M; Cheeramelil, Jacob

2013-12-01

Calretinin a 29-kDa calcium binding protein is expressed widely in normal human tissue and tumours including amelobastoma. The objective of this study was to determine calretinin expression in heamatoxylin and eosin diagnosed cases of ameloblastoma and non-neoplastic odontogenic cysts. The lining epithelium in 3 cases of radicular cysts, 5 cases of odontogenic keratocysts, 5 cases of dentigerous cysts and 11 cases of ameloblastomas were examined for expression of calretinin. No positive epithelial staining was observed in radicular and dentigerous cysts. In comparison, however 100% of cases of ameloblastomas and 40% of cases of odontogenic karatocysts showed positive calretinin expression. Calretinin may be a specific immunohistochemical marker for ameloblastoma. If there is any possible relation between calretinin expression and neural origin of the odontogenic epithelium and its neoplastic transformation and if calretinin could be used as an early marker to predict the tendency of neoplastic change of odontogenic epithelium could be answered through further researches. How to cite this article: D'Silva S, Sumathi MK, Balaji N, Shetty NK, Pramod KM, Cheeramelil J. Evaluation of Calretinin expression in Ameloblastoma and Non-Neoplastic Odontogenic Cysts - An immunohistochemical study. J Int Oral Health 2013; 5(6):42-8 .

Immunohistochemical localization of human papilloma virus in conjunctival neoplasias: A retrospective study

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Sharma, Anjana; Panda, Anita

2007-01-01

Background: The extent of association of human papilloma virus (HPV) in human conjunctival neoplasias has been debated in studies originating from different parts of the world, but no substantial evidence has been generated on Indian subjects. This prompted us to carry out a retrospective study on conjunctival neoplasias diagnosed over the past 12 years. Materials and Methods: Histopathological and immunohistochemical analysis of 65 specimens of ocular neoplasias and 30 normal controls diagnosed between 1991 and 2002 at a tertiary eye care hospital, was undertaken. Formalin-fixed, paraffin-embedded tissues were reviewed for confirming histopathological diagnosis, presence of koilocytosis and changes related to actinic keratosis. Immunohistochemical analysis was done using HPV-specific monoclonal antibodies. Clinicopathological correlation and the association of HPV antigen with the histopathological features were performed. Results: Out of the 65 cases analyzed, 35 were papillomas and 30 were ocular surface squamous neoplasias (OSSN). The mean age was 48 years with a male preponderance. Histologically, koilocytosis was observed in 17.1% of papillomas and 36.6% of OSSN. Actinic keratosis was present in 33% of OSSN. Immunohistochemically 17.1% conjunctival papillomas stained positive for HPV antigen, all cases of OSSN were negative for HPV. There was no correlation between koilocytosis or actinic keratosis and the detection of HPV antigen. Conclusions: The association between HPV and conjunctival neoplasias is variable in different geographical areas and also depends on the methods of detection used. This study warrants the need for applying more advanced techniques at a molecular level to determine the possible etiology of HPV in conjunctival neoplasias among Asian-Indians. PMID:17699945

Merkel cell carcinoma with an unusual immunohistochemical profile

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L. Pilloni

2009-12-01

Full Text Available The clinical and morphological picture of Merkel cell carcinoma (MCC may be rather challenging; therefore, the immunohistochemical profile plays a relevant role in confirming the microscopic diagnosis. A panel of antibodies including cytokeratins 20, 7 and epithelial membrane antigen, and neuronspecific enolase is used in confirming the morphological diagnosis of MCC. The majority of MCCs express CK20 and are CK7-negative. Herein, we present a case of primary cutaneous neuroendocrine carcinoma with an atypical immunohistochemical pattern. A 83-years old woman presented with a painless plaque, red to violaceous in colour, located in the leg. The skin tumor was excided, formalin-fixed and paraffinembedded. Tissue sections were immunostained with a panel of antibodies routinely utilized in complex primary skin tumors for evidencing epithelial and neuroendocrine differentiation of tumor cells. The neuroendocrine differentiation of tumor cells was evidenced by their immunoreactivity for synaptophysin, chromograninA and neuron-specific enolase. Tumor cells also showed diffuse cytoplasmic staining for CK7. No immunoreactivity was detected for CK20 and thyroid transcription factor-1. Our data, together with previous rare reports of CK20-/CK7+ MCCs, lay stress on the importance of routinely utilizing a panel of antibodies in the differential diagnosis of complex primary carcinomas of the skin and may have important implications in expanding the differential diagnosis of skin tumors. In particular, caution should be taken in excluding the diagnosis of MCC only on the basis of the absence of reactivity of tumor cells for CK20, favouring the wrong diagnosis of less aggressive skin tumors.

The clinicopathologic association of c-MET overexpression in Iranian gastric carcinomas; an immunohistochemical study of tissue microarrays

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Sotoudeh Kambiz

2012-05-01

Full Text Available Abstract Background c-MET is an oncogene protein that plays important role in gastric carcinogenesis and has been introduced as a prognostic marker and potential therapeutic target. The aim of this study was to evaluate the frequency of c-MET overexpression and its relationship with clinicopathological variables in gastric cancer of Iranian population using tissue microarray. Methods In a cross sectional study, representative paraffin blocks of 130 patients with gastric carcinoma treated by curative gastrectomy during a 2 years period of 2008–2009 in two university hospitals in Tehran-Iran were collected in tissue microarray and c-MET expression was studied by immunohistochemical staining. Results Finally 124 cases were evaluated, constituted of 99 male and 25 female with the average age of 61.5 years. In 71% (88/124 of tumors, c-MET high expression was found. c-MET high expression was more associated with intestinal than diffuse tumor type (P = 0.04, deeper tumor invasion, pT3 and pT4 versus pT1 and pT2 (P = 0.014, neural invasion (P = 0.002 and advanced TNM staging, stage 3 and 4 versus stage 1 and2 (P = 0.044. The c-MET high expression was not associated with age, sex, tumor location, differentiation grade and distant metastasis, but relative associations with lymph node metastasis (P = 0.065 and vascular invasion (P = 0.078 were observed. Conclusions c-MET oncogene protein was frequently overexpressed in Iranian gastric carcinomas and it was related to clinicopathological characteristics such as tumor type, depth of invasion, neural invasion and TNM staging. It can also support the idea that c-MET is a potential marker for target therapy in Iranian gastric cancer. Virtual slides The virtual slide(s for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9744598757151429

Immunohistochemical Studies of the Kynurenine Pathway in Morphea

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Noakes, Rowland; Mellick, Nick

2013-01-01

Cutaneous sclerosis, resembling that seen in subcutaneous morphea, is a feature of eosinophilic fasciitis and eosinophilia-myalgia syndrome, two conditions in which the kynurenine pathway is known to be activated. To investigate the possibility of activation of the kynurenine pathway in morphea, skin biopsies were taken from involved and non-involved sites in a series of three patients with morphea. Immunohistochemical stains for quinolinic acid and indoleamine 2,3-dioxygenase (IDO) were perf...

High-throughput immunophenotyping of 43 ferret lymphomas using tissue microarray technology

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Hammer, Anne Sofie; Williams, B.; Dietz, H.H.

2007-01-01

To validate the use of the tissue microarray (TMA) method for immunophenotyping of ferret lymphomas, a TMA was constructed containing duplicate 1-mm cores sampled from 112 paraffin-embedded lymphoma tissue specimens obtained from 43 ferret lymphoma cases. Immunohistochemical (IHC) expression of CD3......, CD79 alpha, and Ki-67 (MIB-1) was determined by TMA and whole mount (WM) staining of each individual case for result comparison. There was a high correlation between CD79 alpha and CD3 results comparing ferret TMA and WM sections (kappa statistic 0.71-0.73 for single-core TMA and 0.......79-0.95 for duplicate-core TMA) and between continuous data from Ki-67 staining of ferret TMA sections and WM sections (concordance correlation coefficients 0.77 for single cores and 0.87 for duplicate cores). Subsequently, a panel of commercially available antibodies was applied to the TMA for the analysis...

Immunohistochemical Detection and Localization of Somatostatin Receptor Subtypes in Prostate Tissue from Patients with Bladder Outlet Obstruction

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Rodolfo Montironi

2008-01-01

Full Text Available Background and Aim of the Study: Scant information on the cellular distribution of the five somatostatin receptor (SSTR subtypes in the normal prostate and in neoplasms of the prostate has been reported in very few studies in which techniques, such as in situ hybridization histochemistry, autoradiography, and more recently immunohistochemistry, have been applied. The aim of the study was to examine immunohistochemically the distribution and localization of these 5 subtypes in the various tissue components in normal prostate.

Expression of von Willebrand factor and caldesmon in the placental tissues of pregnancies complicated with intrauterine growth restriction.

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Göksever Çelik, Hale; Uhri, Mehmet; Yildirim, Gökhan

2017-11-02

The decreased placental perfusion is the underlying reason for intrauterine growth restriction that in turn leads to reduced placental perfusion and ischemia. However, there are several issues to be understood in the pathophysiology of intrauterine growth restriction. We aimed to study whether any compensatory response in placental vascular bed occur in pregnancies complicated with intrauterine growth restriction by the immunohistochemical staining of von Willebrand factor and caldesmon in placental tissues. A total of 103 pregnant women was enrolled in the study including 50 patients who were complicated with IUGR and 50 uncomplicated control patients. The study was designed in a prospective manner. All placentas were also stained with von Willebrand factor and caldesmon monoclonal kits. The immunohistochemical staining of von Willebrand factor and caldesmon expressions in placental tissues were different between normal and intrauterine growth restriction group. The percentages of 2+ and 3+ von Willebrand factor expression were higher in the intrauterine growth restriction group comparing with the normal group, although the difference was not statistically significant. The intensity of caldesmon expression was significantly lower in the intrauterine growth restriction group in comparison with the normal group (p intrauterine growth restriction which is a hypoxic condition. But newly formed vessels are immature and not strong enough. Our study is important to clarify the pathophysiology and placental compensatory responses in intrauterine growth restriction.

Immunohistochemical expression of Notch signaling in the lining epithelium of periapical cysts.

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Meliou, Eleni; Kerezoudis, Nikolaos; Tosios, Konstantinos; Lafkas, Daniel; Kiaris, Hippokratis

2011-02-01

In this study we evaluated the immunohistochemical expression of the receptors Notch 1 and Notch 2, the ligand Delta 1, and the transcription factors HES 1 and HES 5 in the epithelium of well-defined periapical cysts. Immunohistochemistry was carried out on 55 formalin-fixed and paraffin-embedded, well-defined periapical cysts with minimum inflammation, obtained from the archival tissue database of the Department of Oral Pathology and Surgery. Western blotting was performed to evaluate the specificity of the anti-Notch antibody and the expression of Notch signaling in 5 fresh-frozen periapical cysts. The levels of staining intensity were estimated by the performance of a semiautomated image analysis system. Descriptive statistic of mean values obtained by computerized image analysis method was performed. Immunostaining reaction of all Notch signaling components was observed in the cytoplasm and/or the cytoplasmic membrane in the majority of epithelial cells of periapical cysts. Nuclear staining was observed occasionally in all cases. Notch 2 showed strong staining in 52.83% of the cases, followed by Notch 1 (35.85%), HES 1 and HES 5 moderate staining in 72.73% and 57.69% of the cases, respectively, and Delta 1 weak staining in 58.33% of the cases. No statistical correlation was found between the antibodies and the sex or the age of the study group. Notch is an evolutionarily conserved signaling mechanism that regulates cell fate decisions during development and postnatal life in organisms as diverse as worms, flies, and humans. The present observations indicate that Notch pathway is active downstream in the lining epithelium of periapical cysts, suggesting an involvement of this pathway in periapical cyst growth and expansion. Copyright © 2011 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Matrix metalloproteinase-1 expression in oral submucous fibrosis: An immunohistochemical study

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Mishra Gauri

2010-01-01

Full Text Available Context: Oral submucous fibrosis (OSF is a form of pathological fibrosis affecting the oral mucosa. There is compelling evidence to implicate the habitual chewing of areca nut with the development of OSF. Because collagens are the major structural components of connective tissues, including oral submucosa, the composition of collagen within each tissue needs to be precisely regulated to maintain tissue integrity. Arecoline stimulates fibroblasts to increase the production of collagen by 150%. Aim: As the role of collagenase is implicated in cleaving the collagen under physical conditions, this study was carried out to evaluate the role of collagenase-1 (matrix metalloproteinase [MMP]-1 in a pathologic condition like OSF. Settings and Design: A total of 40 patients were included in the study, comprising of 30 OSF as Group 1 and 10 normal buccal mucosa tissue as Group 2. Materials and Methods: Both the groups were stained for MMP-1 by the immunohistochemical method using the streptavidin HRP-biotin labeling technique. MMP-1 expression intensity in the epithelium and connective tissue was decreased in Group 1 when compared to Group 2. Statistical Analysis Used: Chi-square test of association was used to determine the difference in the expression of MMP-1 between OSF and normal buccal mucosa and among different histological gradings of OSF. Results: The results were statistically significant. However, there was no statistically significant difference between the expression of MMP-1 among different histological grades of OSF in Group 1.

Comparison of staining of mitotic figures by haematoxylin and eosin-and crystal violet stains, in oral epithelial dysplasia and squamous cell carcinoma

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Ankle Madhuri

2007-01-01

Full Text Available Mitosis of cells gives rise to tissue integrity. Defects during mitosis bring about abnormalities. Excessive proliferation of cells due to increased mitosis is one such outcome, which is the hallmark in precancer and cancer. The localization of proliferating cells or their precursors may not be obvious and easy. Establishing an easy way to distinguish these mitotic cells will help in grading and understanding their biological potential. Although immunohistochemistry is an advanced method in use, the cost and time factor makes it less feasible for many laboratories. Selective histochemical stains like toluidine blue, giemsa and crystal violet have been used in tissues including the developing brain, neural tissue and skin. Aim of the study: 1To compare the staining of mitotic cells in haematoxylin and eosin with that in crystal violet. 2To compare the number of mitotic figures present in normal oral mucosa, epithelial dysplasia and oral squamous cell carcinoma in crystal violet-stained sections with that in H and E-stained sections. Materials and Methods: Ten tissues of normal oral mucosa and 15 tissues each of oral epithelial dysplasia seen in tobacco-associated leukoplakia and squamous cell carcinoma were studied to evaluate the selectivity of 1% crystal violet for mitotic figures. The staining was compared with standard H and E staining. Statistical analysis was done using Man-Whitney U test. Results: A statistically significant increase in the mean mitotic count was observed in crystal violet-stained sections of epithelial dysplasia as compared to the H and E-stained sections ( p = 0.0327. A similar increase in the mitotic counts was noted in crystal violet-stained sections of oral squamous cell carcinoma as compared to the H and E-stained sections.( p = 0.0443. No significant difference was found in the mitotic counts determined in dysplasia or carcinoma by either the crystal violet ( p = 0.4429 or the H and E-staining techniques ( p = 0

Evaluation of cell proliferative activity after irradiation using immunohistochemical approach and flow cytometry

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Tamada, Takashi (Okayama Univ. (Japan). School of Medicine)

1992-06-01

To evaluate a proliferative activity of post-irradiated malignant cells, we studied the kinetics of HeLa cells using immunohistochemical approach and flow cytometry. HeLa cells were stained with two proliferation-associated monoclonal antibodies, Ki-67 and anti-DNA polymerase {alpha} antibody. Nucleoli of non-irradiated cells were granularly stained with Ki-67. After irradiation, only the center of nuclei was diffusely stained with Ki-67. One hundred forty-four hours after low-dose irradiation, the staining patterns became the same as the control. On the other hand, after high-dose irradiation, the center of nuclei was weakly stained. DNA polymerase {alpha} was diffusely labelled with nuclei of the control. It was located around the border of nuclei of low-dose irradiated cells like a ring. But after high-dose irradiation, it was granularly distributed in the periphery of nuclei. FITC conjugated Ki-67/PI two parameter analysis was done by a single laser flow cytometer. Twenty-four hours after irradiation, DNA-histograms showed the accumulation to G{sub 2}/M phase and the increase of DNA content of G{sub 2}/M cells, as exposure dose was increased. Two parameter analysis showed the increase of FITC uptake of G{sub 2}/M phase as dose increased. These changes of flow cytometry were remarkably observed after 24 hours' incubation. It was shown that the difference of Ki-67 antigen and DNA polymerase {alpha} appearance depended on the irradiation dose. These findings suggest that immunohistochemical staining with Ki-67 or anti-DNA polymerase {alpha} antibody and flow cytometry using Ki-67 are available to evaluate cell damages after irradiation. (author).

Evidence for the ectopic synthesis of melanin in human adipose tissue.

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Randhawa, Manpreet; Huff, Tom; Valencia, Julio C; Younossi, Zobair; Chandhoke, Vikas; Hearing, Vincent J; Baranova, Ancha

2009-03-01

Melanin is a common pigment in animals. In humans, melanin is produced in melanocytes, in retinal pigment epithelium (RPE) cells, in the inner ear, and in the central nervous system. Previously, we noted that human adipose tissue expresses several melanogenesis-related genes. In the current study, we confirmed the expression of melanogenesis-related mRNAs and proteins in human adipose tissue using real-time polymerase chain reaction and immunohistochemical staining. TYR mRNA signals were also detected by in situ hybridization in visceral adipocytes. The presence of melanin in human adipose tissue was revealed both by Fontana-Masson staining and by permanganate degradation of melanin coupled with liquid chromatography/ultraviolet/mass spectrometry determination of the pyrrole-2,3,5-tricarboxylic acid (PTCA) derivative of melanin. We also compared melanogenic activities in adipose tissues and in other human tissues using the L-[U-(14)C] tyrosine assay. A marked heterogeneity in the melanogenic activities of individual adipose tissue extracts was noted. We hypothesize that the ectopic synthesis of melanin in obese adipose may serve as a compensatory mechanism that uses its anti-inflammatory and its oxidative damage-absorbing properties. In conclusion, our study demonstrates for the first time that the melanin biosynthesis pathway is functional in adipose tissue.

Histomorphometric and immunohistochemical analysis of human maxillary sinus-floor augmentation using porous β-tricalcium phosphate for dental implant treatment.

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Miyamoto, Shinji; Shinmyouzu, Kouhei; Miyamoto, Ikuya; Takeshita, Kenji; Terada, Toshihisa; Takahashi, Tetsu

2013-08-01

This study utilized the constitution and expression of Runx2/Cbfa1 to conduct 6-month-post-operation histomorphometrical and histochemical analysis of osteocalcin in bone regeneration following sinus-floor augmentation procedures using β-tricalcium phosphate (β-TCP) and autogenous cortical bone. Thirteen sinuses of nine patients were treated with sinus-floor augmentation using 50% β-TCP and 50% autogenous cancellous bone harvested from the ramus of the mandible. Biopsies of augmented sinuses were taken at 6 months for histomorphometric and immunohistochemical measurements. Runx2/Cbfa1- and osteocalcin-positive cells were found around TCP particles and on the bone surface. Approximately 60% of cells found around TCP particles stained positive for Runx2/Cbfa1. Fewer cells stained positive for osteocalcin. These positive cells decreased apically with increasing vertical distance from the maxillary bone surface. Histomorphometric analysis showed that the augmented site close to residual bone and periosteum contained approximately 42% bony tissue and 42% soft connective tissue, and the remaining 16% consisted of TCP particles. On the other hand, the augmented bone far from residual bone and periosteum contained 35% bony tissue and 50% soft connective tissue. Our data suggest that TCP particles attract osteoprogenitor cells that migrate into the interconnecting micropores of the bone-substitute material by 6 months. The augmented site close to residual bone contained a higher proportion of bony tissue and a lower proportion of soft connective tissue than did the augmented site far from residual bone. © 2012 John Wiley & Sons A/S.

Double immunohistochemical staining with laminin 5 (γ2 chain) and collagen IV in colorectal neoplasms

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Fiehn, Anne-Marie Kanstrup; Bzorek, Michael; Warnecke, Mads

2016-01-01

divided according to the primary histopathological diagnoses of tubular adenoma, tubulovillous adenoma, adenoma with pseudoinvasion and glandular adenocarcinoma stages pT1, pT2 or pT3, were included in the study. In normal colonic mucosa, no expression of laminin 5 staining was observed. BM was always...... as a supplement for the diagnosis of pT1 CRC. In adenomas, the double staining highlights the areas for the pathologist to pay extra attention. By itself, the double staining cannot determine whether or not there is invasion. Morphology remains the single most important factor in differentiating adenoma...

Immunohistochemical identification of messenger RNA-related proteins in basophilic inclusions of adult-onset atypical motor neuron disease.

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Fujita, Kengo; Ito, Hidefumi; Nakano, Satoshi; Kinoshita, Yoshimi; Wate, Reika; Kusaka, Hirofumi

2008-10-01

This report concerns an immunohistochemical investigation on RNA-related proteins in the basophilic inclusions (BIs) from patients with adult-onset atypical motor neuron disease. Formalin-fixed, paraffin-embedded sections of the motor cortex and the lumbar spinal cord were examined. The BIs appeared blue in color with H&E and Nissl stain, and pink with methylgreen-pyronin stain. Ribonuclease pretreatment abolished the methylgreen-pyronin staining, suggesting that the BIs contained RNA. Immunohistochemically, the BIs were distinctly labeled with the antibodies against poly(A)-binding protein 1, T cell intracellular antigen 1, and ribosomal protein S6. These proteins are essential constituents of stress granules. In contrast, the BIs were not immunoreactive for ribosomal protein L28 and decapping enzyme 1, which are core components of transport ribonucleoprotein particles and processing bodies, respectively. Moreover, the BIs were not immunopositive for TDP-43. Our results imply that translation attenuation could be involved in the processes of BI formation in this disorder.

Immunohistochemical Studies of the Kynurenine Pathway in Morphea

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Rowland Noakes

2013-01-01

Full Text Available Cutaneous sclerosis, resembling that seen in subcutaneous morphea, is a feature of eosinophilic fasciitis and eosinophilia-myalgia syndrome, two conditions in which the kynurenine pathway is known to be activated. To investigate the possibility of activation of the kynurenine pathway in morphea, skin biopsies were taken from involved and non-involved sites in a series of three patients with morphea. Immunohistochemical stains for quinolinic acid and indoleamine 2,3-dioxygenase (IDO were performed.

Ocular melanoma metastatic to skin: the value of HMB-45 staining.

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Schwartz, Robert A; Kist, Joseph M; Thomas, Isabelle; Fernández, Geover; Cruz, Manuel A; Koziorynska, Ewa I; Lambert, W Clark

2004-06-01

Cutaneous metastatic disease is an important finding that may represent the first sign of systemic cancer, or, if already known, that may change tumor staging and thus dramatically altered therapeutic plans. Although cutaneous metastases are relatively frequent in patients with cutaneous melanoma, they are less so from ocular melanoma. To demonstrate the value of HMB-45, staining in the detection of ocular melanoma metastatic to skin. The immunohistochemical stain HMB-45 a monoclonal antibody directed against intact human melanoma cells, was employed on a skin biopsy specimen from a cutaneous tumor. HMB-45 staining was positive in the atypical hyperchromatic cells of the deep dermis. HMB-45 may be of value in the detection of ocular melanoma metastatic to skin. Cutaneous metastatic disease is a somewhat common and extremely important diagnosis. Although cutaneous metastases from cutaneous melanoma are relatively frequent, those from ocular melanomas are less so. Use of histochemical staining, especially the HMB-45 stain, allows confirmation of the diagnosis.

Evaluation of several immunohistochemical markers in malignant mesothelioma and pulmonary adenocarcinoma

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Zhao Xiaoguang

1992-01-01

The role of CEA, keratin, vimentin and secretary component (SC) in the diagnosis of malignant mesothelioma (MM) and pulmonary adenocarcinoma (PA) was evaluated by immunohistochemical method. 30 and 22 PA exhibited positive reactions for CEA and SC respectively. In contrast, all MM (8 cases) failed to stain for CEA and SC. We conclude that use of anti-keratin serum can not provide a help for the differential diagnosis of MM and PA. Positive reaction for CEA and SC favours a diagnosis of PA, while positive staining for vimentin but negative for CEA and SC favours the diagnosis of MM

Immunohistochemical study of the expression of cell cycle regulating proteins at different stages of bladder cancer

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Primdahl, Hanne; von der Maase, Hans; Sørensen, Flemming Brandt

2002-01-01

; group 1, 23 patients with recurrent stage Ta (non-invasive) tumors; group 2, 22 patients presenting at their first admission with T2-4 (muscle invasive) tumors; group 3, 24 patients who experienced disease progression from Ta or T1 (invasive in connective tissue) to a higher stage......PURPOSE: The cell cycle is known to be deregulated in cancer. We therefore analyzed the expression of the cell cycle related proteins p21, p27, p16, Rb, and L-myc by immunohistochemical staining of bladder tumors.METHODS: The tissue material consisted of bladder tumors from three groups of patients......(kip1) ( P=0.03), Rb ( P=0.00002), and L-myc ( P=0.00000007) in muscle invasive tumors compared to noninvasive tumors. Tumors presenting as muscle invasive at first diagnosis had significantly lower levels of p16/CDKN2A ( P=0.01) when compared to muscle invasive tumors that followed Ta or T1 precursor...

Immunohistochemical correlation of matrix metalloproteinase-2 and tissue inhibitors of metalloproteinase-2 in tobacco associated epithelial dysplasia.

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Bajracharya, Dipshikha; Shrestha, Bijayatha; Kamath, Asha; Menon, Aparna; Radhakrishnan, Raghu

2014-01-01

To study the immunohistochemical expression of matrix metalloproteinase and tissue inhibitors of matrix metalloproteinase-2 in different histological grades of tobacco associated epithelial dysplasia and correlate the association between these proteases. Potentially malignant oral disorders (PMODs) progressing to oral cancer are related to the severity of epithelial dysplasia. A retrospective immunohistochemical study was carried out on 30 clinically and histologically proven cases of leukoplakia with dysplasia and 10 cases of normal buccal mucosa using anti-MMP-2 and anti-TIMP-2 monoclonal antibodies. Mann Whitney U test, for comparing the expression of both MMP-2 and TIMP-2 in normal mucosa with dysplasia, was highly significant (P correlation between MMP-2 and TIMP-2 through different grades of dysplasia and cells observed showed positive correlation. Concomitant increase in the expression of both MMP-2 and TIMP-2 suggested that the activation of MMP-2 is dependent on TIMP-2 acting as a cofactor. Changes in TIMP-2 levels are considered important because they directly affect the level of MMP-2 activity.

Levey-Jennings Analysis Uncovers Unsuspected Causes of Immunohistochemistry Stain Variability.

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Vani, Kodela; Sompuram, Seshi R; Naber, Stephen P; Goldsmith, Jeffrey D; Fulton, Regan; Bogen, Steven A

Almost all clinical laboratory tests use objective, quantitative measures of quality control (QC), incorporating Levey-Jennings analysis and Westgard rules. Clinical immunohistochemistry (IHC) testing, in contrast, relies on subjective, qualitative QC review. The consequences of using Levey-Jennings analysis for QC assessment in clinical IHC testing are not known. To investigate this question, we conducted a 1- to 2-month pilot test wherein the QC for either human epidermal growth factor receptor 2 (HER-2) or progesterone receptor (PR) in 3 clinical IHC laboratories was quantified and analyzed with Levey-Jennings graphs. Moreover, conventional tissue controls were supplemented with a new QC comprised of HER-2 or PR peptide antigens coupled onto 8 μm glass beads. At institution 1, this more stringent analysis identified a decrease in the HER-2 tissue control that had escaped notice by subjective evaluation. The decrement was due to heterogeneity in the tissue control itself. At institution 2, we identified a 1-day sudden drop in the PR tissue control, also undetected by subjective evaluation, due to counterstain variability. At institution 3, a QC shift was identified, but only with 1 of 2 controls mounted on each slide. The QC shift was due to use of the instrument's selective reagent drop zones dispense feature. None of these events affected patient diagnoses. These case examples illustrate that subjective QC evaluation of tissue controls can detect gross assay failure but not subtle changes. The fact that QC issues arose from each site, and in only a pilot study, suggests that immunohistochemical stain variability may be an underappreciated problem.

Immunohistochemical findings of the granulomatous reaction associated with tuberculosis

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Shirin Karimi

2016-01-01

Conclusion: Detection of TB in tissue slides is still based on the histological pattern of the granuloma, which has several differential diagnoses with different treatments. Presence of mycobacterial antigens and tissue morphology can be evaluated using the IHC technique. Considering the criteria of positive IHC staining of TB granulomatous reactions, this stain not only highlights the presence of mycobacterial antigens for tissue diagnosis, but also could morphologically localize their distribution in different cells. Pathologists must be familiar with adequate staining pattern, elimination of background staining, and type of selected antibody. This method is especially important for application in countries with high prevalence of TB as a technique with early diagnostic value in tissue specimens. Early diagnosis using this technique can reduce related morbidity and mortality and decrease the rate of complications due to misdiagnosis and mistreatment of TB.

Morphological and immunohistochemical criteria of tissue response to radiotherapy in rectal cancer.

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Ionescu, S; Brătucu, E; Zurac, S; Staniceanu, F; Pătraşcu, Tr; Burcoş, Tr; Herlea, V; Degeratu, D; Popa, I; Cristian, D

2013-01-01

Given the context that rectal tumours respond to a certain degree to radiotherapy, a necessity arises for estimating a tumour's capacity to react to radiation from the very moment of diagnostic biopsy. We have histologically and immunohistochemically analysed tissues coming from 52 patients with rectal adenocarcinomas. Of the studied parameters, the ones presenting significant variation under radiotherapy in terms of statistics(p 0.05) were: colloid type (p=0.001), EGFR in the tumour(p=0.00045), EGFR in the normal epithelium (p=0.0017),VEGF in the tumour (p=0.0132) and VEGF in the tumour stroma (p=0.030). Our study follows the same trends as the medical literature we have consulted regarding the variation of EGFR and VEGF with radiotherapy, and the distinct note of our study relies in the observation that normal stroma in case of rectal tumors also reacts to radiotherapy, sometimes more aggressively than the tumor itself, especially in which concerns the nerve and muscle fibers. Celsius.

Development of murine monoclonal antibodies for the immunohistochemical diagnosis of systemic bovine aspergillosis

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Jensen, H.E.; Aalbaek, B.; Lind, Peter

1996-01-01

) in immunohistochemical and immunoblotting assays. In immunohistochemical assays, all MAbs raised against WSSA cross-reacted heavily with a number of other fungal species. All 4 MAbs (MAb-WF-AF-1-4) raised against the WF reacted strongly with hyphae of Aspergillus spp.; hyphae of Scedosporium apiospermum were also......), the MAb-WF-AF-1 and the polyclonal anti-Aspergillus antibodies reacted in a similar pattern, i.e., positively in 41 aspergillosis lesions and negatively in 92 zygomycotic lesions. Hyphae in 3 of 12 lesions that were not stained by the polyclonal antibodies reacted with the specific MAb-WF-AF-1; i...

Immunohistochemical expression of Insulin-like growth factor-1, Transforming growth factor-beta1, and Vascular endothelial growth factor in parathyroid adenoma and hyperplasia

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Hamide Sayar

2014-01-01

Full Text Available Background: Insulin-like growth factor (IGF, transforming growth factor-beta1 (TGF-β1, and vascular endothelial growth factor (VEGF are commonly studied growth factors, but little data are available on the immunohistochemical expression of these factors in parathyroid lesions. Materials and Methods: Tissue specimens from 36 patients with primary hyperparathyroidism (P-HPT (26 adenomas and 10 primary hyperplasias were examined. Normal parathyroid tissue adjacent to the adenoma or area of hyperplasia was used as control tissue. Preoperative laboratory testing [serum Ca and P, creatinine and parathormone levels (PTH] which led to the diagnosis of P-HPT had been performed, the size and weight of the parathyroid glands measured, and postoperative serum PTH levels determined. Paraffin-embedded parathyroid tissue specimens were stained with antibodies to IGF-1, VEGF, and TGF-β1 using standard immunohistochemical procedures. Results: IGF-1 immunoreactivity was seen in 50% of hyperplasia and in 46% of adenoma samples, but in 87% of normal parathyroid tissue in the vicinity of the adenomas (P = 0.005. TGF-β1 immunoreactivity was observed in 90% of hyperplasia, in 92% of adenoma samples, and in 95% of normal tissues around adenomas. VEGF immunoreactivity was observed in 70% of hyperplastic and 65% of adenomatous tissues, as well as in 54% of normal tissues in the vicinity of the adenoma. No significant differences in the expression of IGF-1, TGF-β1, and VEGF were observed between primary adenomas compared to hyperplasia samples (P > 0.05. Conclusions: Parathyroid tissue is clearly a site for production of IGF-1, TGF-β1, and VEGF. IGF-1 receptor activity was higher in normal parathyroid tissue compared to hyperplastic and adenomatous tissue.

Histopathologic and immunohistochemical features of capsular tissue around failed Ahmed glaucoma valves.

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Alka Mahale

Full Text Available Impervious encapsulation around Ahmed glaucoma valve (AGV results in surgical failure raising intraocular pressure (IOP. Dysregulation of extracellular matrix (ECM molecules and cellular factors might contribute to increased hydraulic resistance to aqueous drainage. Therefore, we examined these molecules in failed AGV capsular tissue. Immunostaining for ECM molecules (collagen I, collagen III, decorin, lumican, chondroitin sulfate, aggrecan and keratan sulfate and cellular factors (αSMA and TGFβ was performed on excised capsules from failed AGVs and control tenon's tissue. Staining intensity of ECM molecules was assessed using Image J. Cellular factors were assessed based on positive cell counts. Histopathologically two distinct layers were visible in capsules. The inner layer (proximal to the AGV showed significant decrease in most ECM molecules compared to outer layer. Furthermore, collagen III (p = 0.004, decorin (p = 0.02, lumican (p = 0.01 and chondroitin sulfate (p = 0.02 was significantly less in inner layer compared to tenon's tissue. Outer layer labelling however was similar to control tenon's for most ECM molecules. Significantly increased cellular expression of αSMA (p = 0.02 and TGFβ (p = 0.008 was detected within capsular tissue compared to controls. Our results suggest profibrotic activity indicated by increased αSMA and TGFβ expression and decreased expression of proteoglycan (decorin and lumican and glycosaminoglycans (chondroitin sulfate. Additionally, we observed decreased collagen III which might reflect increased myofibroblast contractility when coupled with increased TGFβ and αSMA expression. Together these events lead to tissue dysfunction potentially resulting in hydraulic resistance that may affect aqueous flow through the capsular wall.

Histopathologic and immunohistochemical features of capsular tissue around failed Ahmed glaucoma valves.

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Mahale, Alka; Fikri, Fatma; Al Hati, Khitam; Al Shahwan, Sami; Al Jadaan, Ibrahim; Al Katan, Hind; Khandekar, Rajiv; Maktabi, Azza; Edward, Deepak P

2017-01-01

Impervious encapsulation around Ahmed glaucoma valve (AGV) results in surgical failure raising intraocular pressure (IOP). Dysregulation of extracellular matrix (ECM) molecules and cellular factors might contribute to increased hydraulic resistance to aqueous drainage. Therefore, we examined these molecules in failed AGV capsular tissue. Immunostaining for ECM molecules (collagen I, collagen III, decorin, lumican, chondroitin sulfate, aggrecan and keratan sulfate) and cellular factors (αSMA and TGFβ) was performed on excised capsules from failed AGVs and control tenon's tissue. Staining intensity of ECM molecules was assessed using Image J. Cellular factors were assessed based on positive cell counts. Histopathologically two distinct layers were visible in capsules. The inner layer (proximal to the AGV) showed significant decrease in most ECM molecules compared to outer layer. Furthermore, collagen III (p = 0.004), decorin (p = 0.02), lumican (p = 0.01) and chondroitin sulfate (p = 0.02) was significantly less in inner layer compared to tenon's tissue. Outer layer labelling however was similar to control tenon's for most ECM molecules. Significantly increased cellular expression of αSMA (p = 0.02) and TGFβ (p = 0.008) was detected within capsular tissue compared to controls. Our results suggest profibrotic activity indicated by increased αSMA and TGFβ expression and decreased expression of proteoglycan (decorin and lumican) and glycosaminoglycans (chondroitin sulfate). Additionally, we observed decreased collagen III which might reflect increased myofibroblast contractility when coupled with increased TGFβ and αSMA expression. Together these events lead to tissue dysfunction potentially resulting in hydraulic resistance that may affect aqueous flow through the capsular wall.

LANTHANUM STAINING OF THE SURFACE COAT OF CELLS

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Shea, Stephen M.

1971-01-01

Among the techniques which have been reported to stain the surface coat of cells, for electron microscopy, is lanthanum staining en bloc. Similarly, the presence of the cationic dye, Alcian blue 8GX, in a primary glutaraldehyde fixative has been reported to improve the preservation of the surface coat of cells of many types; however, the preserved coat is not very electron opaque unless thin sections are counterstained. The present paper shows that for several rat tissues lanthanum staining en bloc is an effective electron stain for the cell surface, giving excellent contrast, if combined sequentially with prefixation in an aldehyde fixative containing Alcian blue. The cationic substance cetylpyridinium chloride was found to have a similar effect to that of Alcian blue in enhancing the lanthanum staining of the surface coat material of the brush border of intestinal epithelial cells. The patterns of lanthanum staining obtained for the tissues studied strikingly resemble those reported in the literature where tissues are stained by several standard methods for demonstrating mucosubstances at the ultrastructural level. This fact and the reproduction of the effect of Alcian blue by cetylpyridinium chloride constitute a persuasive empirical argument that the material visualized is a mucopolysaccharide or mucopolysaccharide-protein complex. PMID:4108476

Histological Stains: A Literature Review and Case Study.

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Alturkistani, Hani A; Tashkandi, Faris M; Mohammedsaleh, Zuhair M

2015-06-25

The history of histology indicates that there have been significant changes in the techniques used for histological staining through chemical, molecular biology assays and immunological techniques, collectively referred to as histochemistry. Early histologists used the readily available chemicals to prepare tissues for microscopic studies; these laboratory chemicals were potassium dichromate, alcohol and the mercuric chloride to harden cellular tissues. Staining techniques used were carmine, silver nitrate, Giemsa, Trichrome Stains, Gram Stain and Hematoxylin among others. The purpose of this research was to assess past and current literature reviews, as well as case studies, with the aim of informing ways in which histological stains have been improved in the modern age. Results from the literature review has indicated that there has been an improvement in histopathology and histotechnology in stains used. There has been a rising need for efficient, accurate and less complex staining procedures. Many stain procedures are still in use today, and many others have been replaced with new immunostaining, molecular, non-culture and other advanced staining techniques. Some staining methods have been abandoned because the chemicals required have been medically proven to be toxic. The case studies indicated that in modern histology a combination of different stain techniques are used to enhance the effectiveness of the staining process. Currently, improved histological stains, have been modified and combined with other stains to improve their effectiveness.

Morphological and immunohistochemical characterisation of seminomas in Norwegian dogs

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Thorvaldsen Tor

2012-09-01

Full Text Available Abstract Background Seminomas in the dog have traditionally been assumed to resemble human spermatocytic seminomas, based on their low malignancy and high occurrence in old individuals. However, recently published studies indicate that canine seminomas can be classified as classical and spermatocytic seminomas in a similar way as in man, and that classical seminomas comprise a substantial proportion of seminomas in the dog. These two factors both contribute to increasing the potential of canine seminoma as a relevant model for human testicular cancer. The aim of the present study was to characterise seminoma in Norwegian dogs using morphology and immunohistochemistry, and determine whether these tumours are comparable with human classical seminoma. Methods By applying diagnostic criteria from human pathology, 45 seminomas from the Norwegian Canine Cancer Register were examined histologically with hematoxylin and eosin (HE and periodic acid-Schiff (PAS stains. All sections were stained immunohistochemically with antibodies against human placental alkaline phosphatase (PLAP and the transmembrane receptor c-KIT. Results Although two of the seminomas showed immunohistochemical staining characteristics indicative of classical seminoma (PLAP+/c-KIT+, all 45 examined seminomas were morphologically consistent with spermatocytic seminoma. Conclusions The value of canine seminoma as a model for SE in man remains unclear. Among the 45 investigated tumours from Norwegian dogs, none were classified as classical seminoma based on morphological criteria consistent with human seminomas. Regional or breed differences in the occurrence of classical seminoma in the dog, as well as the lack of uniform diagnostic criteria, might explain the discrepancy between the findings in the current study and the results presented by other authors.

Ductal adenocarcinoma of the prostate: immunohistochemical findings and clinical significance

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Sha JJ

2013-10-01

Full Text Available Jianjun Sha,1,2 Juanjie Bo,1 Jiahua Pan,1 Lianhua Zhang,1 Hanqing Xuan,1 Wei Chen,1 Dong Li,1 Zhaoliang Wang,1 Dongming Liu,1 Yiran Huang1,2 1Department of Urology, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, 2School of Biomedical Engineering, Shanghai Jiaotong University, Shanghai, People's Republic of China Introduction: To investigate the clinical features, diagnosis, treatment, and prognosis of ductal adenocarcinoma of the prostate. Methods: The clinicopathological and immunohistochemical data of seven patients with ductal adenocarcinoma of the prostate were retrospectively analyzed. All patients underwent physical examination, magnetic resonance imaging (MRI, bone scan, cystoscopy, and computed tomography (CT scan. The level of prostate-specific antigen (PSA before and after surgery was assessed. Different prostate cancer markers were used for immunohistochemical staining. Results: The mean age of the seven patients diagnosed with prostatic ductal adenocarcinoma in this study was 76.2 years (range 57–88. Five patients presented with intermittent and painless gross hematuria, one patient with progressive dysuria, and one patient with elevated serum PSA on routine health examination. The level of PSA before surgery ranged from 1.3 to 45.0 ng/mL. Immunohistochemical staining results of the prostatic ductal adenocarcinoma confirmed positivity for PSA, prostatic acid phosphatase, androgen receptor, and alpha-methyacyl co-enzyme A (CoA-reductase markers. Two of the patients underwent bilateral orchiectomy combined with anti-androgen therapy, three underwent transurethral resection of prostate, one received radical prostatectomy, and one received medical castration therapy. The clinical outcomes of all patients were satisfactory, based on follow-up data. The symptoms of hematuria and dysuria were ameliorated well, and the postoperative PSA level decreased below 4.0 ng/mL. Recurrence or metastasis of disease was

Immunohistochemical detection of SWC3, CD2, CD3, CD4 and CD8 antigens in paraformaldehyde fixed and paraffin embedded porcine lymphoid tissue

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Tingstedt, Jens Erik; Tornehave, Ditte; Lind, Peter

2003-01-01

Identification of the different cell types of the immune system is important for in situ studies on the pathogenesis of infectious diseases in various animals, including the pig. Unfortunately, many monoclonal anti-leukocyte antibodies are only useful for staining frozen tissue sections with inhe......Identification of the different cell types of the immune system is important for in situ studies on the pathogenesis of infectious diseases in various animals, including the pig. Unfortunately, many monoclonal anti-leukocyte antibodies are only useful for staining frozen tissue sections...... with inherent poor tissue morphology, and are not readily adapted to formaldehyde fixed and paraffin embedded tissue with well preserved morphology. Seven well characterised monoclonal antibodies against porcine leukocyte antigens were tested on neutral buffered paraformaldehyde fixed and paraffin embedded...

Immunohistochemical localization of epidermal growth factor in rat and man

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Poulsen, Steen Seier; Nexø, Ebba

1986-01-01

Epidermal growth factor (EGF) is a peptide which stimulates cell mitotic activity and differentiation, has a cytoprotective effect on the gastroduodenal mucosa, and inhibits gastric acid secretion. The immunohistochemical localization of EGF in the Brunner's glands and the submandibular glands is...... antisera against human urinary EGF worked in rat as well as man. EGF was found only in cells with an exocrine function.......Epidermal growth factor (EGF) is a peptide which stimulates cell mitotic activity and differentiation, has a cytoprotective effect on the gastroduodenal mucosa, and inhibits gastric acid secretion. The immunohistochemical localization of EGF in the Brunner's glands and the submandibular glands...... is well documented. The localization of EGF in other tissues is still unclarified. In the present study, the immunohistochemical localization of EGF in tissues from rat, man and a 20 week human fetus were investigated. In man and rat, immunoreaction was found in the submandibular glands, the serous glands...

Immunohistochemical differentiation of atypical hyperplasia vs. carcinoma in situ of the breast.

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Masood, S; Sim, S J; Lu, L

1992-01-01

The distinction between atypical hyperplasia and carcinoma in situ in breast lesions can be difficult. The identification of myoepithelial cell layers may be helpful in establishing a diagnosis of proliferative breast disease vs. intraepithelial neoplasia. We reviewed pathologic material on 20 cases of atypical hyperplasia and 29 cases of carcinoma in situ. Immunohistochemical stains were employed against muscle-specific actin, S-100 protein, and cytokeratin to identify myoepithelial cells and to recognize different staining patterns. In atypical hyperplasia, muscle-specific actin staining identified myoepithelial cells in fine branching fibrovascular layers or as scattered cells between other proliferating cells. This pattern was absent in carcinoma in situ. S-100 protein showed more positive staining in atypical hyperplasia than in carcinoma in situ with patterns distinct from muscle-specific actin. Immunostaining for cytokeratin demonstrated distinctly different patterns between the two lesions. This study suggests that muscle-specific actin, S-100 protein, and cytokeratin in combination may assist in distinguishing proliferative breast disease with atypia from carcinoma in situ.

Immunohistochemical loss of 5-hydroxymethylcytosine expression in acute myeloid leukaemia: relationship to somatic gene mutations affecting epigenetic pathways.

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Magotra, Minoti; Sakhdari, Ali; Lee, Paul J; Tomaszewicz, Keith; Dresser, Karen; Hutchinson, Lloyd M; Woda, Bruce A; Chen, Benjamin J

2016-12-01

Genes affecting epigenetic pathways are frequently mutated in myeloid malignancies, including acute myeloid leukaemia (AML). The genes encoding TET2, IDH1 and IDH2 are among the most commonly mutated genes, and cause defective conversion of 5-methylcytosine into 5-hydroxymethylcytosine (5hmC), impairing demethylation of DNA, and presumably serving as driver mutations in leukaemogenesis. The aim of this study was to correlate 5hmC immunohistochemical loss with the mutation status of genes involved in epigenetic pathways in AML. Immunohistochemical staining with an anti-5hmC antibody was performed on 41 decalcified, formalin-fixed paraffin-embedded (FFPE) bone marrow biopsies from patients with AML. Archived DNA was subjected to next-generation sequencing for analysis of a panel of genes, including TET2, IDH1, IDH2, WT1 and DNMT3A. TET2, IDH1, IDH2, WT1 and DNMT3A mutations were found in 46% (19/41) of the cases. Ten of 15 cases (67%) with TET2, IDH1, IDH2 or WT1 mutations showed deficient 5hmC staining, whereas nine of 26 cases (35%) without a mutation in these genes showed loss of 5hmC. It is of note that all four cases with TET2 mutations showed deficient 5hmC staining. Overall, somatic mutations in TET2, IDH1, IDH2, WT1 and DNMT3A were common in our cohort of AML cases. Immunohistochemical staining for 5hmC was lost in the majority of cases harbouring mutations in these genes, reflecting the proposed relationship between dysfunctional epigenetic pathways and leukaemogenesis. © 2016 John Wiley & Sons Ltd.

Immunohistochemical expression of HER-2/neu in patients with lung carcinoma and its prognostic significance

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Petrusevska, G.; Banev, S.; Ilievska-Poposka, B.; Smickova, S.; Spirovski, Z.

2004-01-01

Background. The HER-2 protein or p185her2 is a membrane receptor with tyrosine kinase activity encoded by HER-2/neu gene. Overexpression of HER-2/neu has been observed in many human cancers, including lung cancer. In the study, the expression of HER-2 protein is determined in the spectrum of lung cancer (adenocarcinoma, squamous cell carcinoma and small cell carcinoma). Patients and methods. The study population consisted of two groups: 19 patients that had undergone surgical treatment and 10 patients that had undergone fiber-optic bronchoscopy and biopsy for primary diagnosis only. Tissue specimens were neutral formaldehyde-fixed and paraffin-embedded. Standard histochemical and immunohistochemical staining were used for diagnosis. Expression of HER-2/neu protein was determined by immunohistochemical staining with Hercep Test (DAKO). The results were graded 0-1 as negative and 2-3 as positive. Results. Overall incidence of HER-2/neu overexpression was 34.4% (10 of 29). Higher incidence was found in the patients with adenocarcinoma 45.4% (5 of 11). In squamous cell carcinoma and small cell carcinoma, the overexpression incidence was 30.7% (4 of 13) and 20% (1 of 5), respectively. No statistically significant difference was seen given the age and gender. HER-2/neu overexpression was more pronounced in the patients with advanced tumour: all patients with squamous cell carcinoma and HER-2/neu overexpression had stage IIIB and stage IV disease, while 80 % of adenocarcinoma patients with HER-2/neu overexpression had stage IIIA and IIIB disease. Conclusions. These results are satisfactory and encourage us to continue this work in the follow-up study to evaluate HER-2/neu role as predictive and prognostic factor for the patients with lung cancer. (author)

Immunohistochemical expression of basement membrane proteins of verrucous carcinoma of the oral mucosa.

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Arduino, Paolo G; Carrozzo, Marco; Pagano, Marco; Broccoletti, Roberto; Scully, Crispian; Gandolfo, Sergio

2010-06-01

Squamous cell carcinoma (SCC) of the oral cavity is an extremely invasive tumour of stratified squamous epithelium that spreads throughout degradation of the basement membrane (BM) and extra-cellular matrix. Oral verrucous carcinoma (VC) is a rare low-grade variant of oral SCC that penetrates into the subepithelial connective tissue. It also has a different clinical behaviour from classical oral SCC. We investigated the immunohistochemical expression of laminin, laminin-5, collagen IV and fibronectin in VC, severe epithelial dysplasia (SED) and SCC in order to analyse if the pattern of these molecules expression contributes to the differences in the biological behaviour of these diseases. The staining pattern of laminin was less intensive in SCC compared with SED and VC, and collagen IV expression was increased in VC compared with SED. Discontinuities of laminin, collagen IV and fibronectin were more evident in SED than in VC. This study indicates that VC has a biological behaviour different from SED or SCC, observable by immunohistochemistry in the BM zone.

Immunohistochemical localization of integrin alpha V beta 3 and osteopontin suggests that they do not interact during embryo implantation in ruminants

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MacLaren Leslie A

2004-04-01

Full Text Available Abstract Background It has been suggested that trophoblast attachment requires co-expression of integrin alpha V beta 3 and its ligand osteopontin at the fetal-maternal interface. Until now the expression patterns of integrin alpha V beta 3 and osteopontin in the pregnant bovine uterus were unknown. The objectives of this study were to localize integrin alpha V beta 3 and osteopontin in bovine and sheep endometrium during the periimplantation period and to compare the distribution patterns using antibodies that had not yet been tested in sheep. Methods Cell compartments within endometrial tissue sections were scored for immunohistochemical staining intensity and data were analyzed to determine the effects of day of pregnancy or cycle. Results In pregnant bovine endometrium, integrin alpha V beta 3 was detected in luminal epithelium, stroma, myometrium and smooth muscle. A strong band of immunoreactivity was observed in the subepithelial stroma of intercaruncular regions, but there was reduced reactivity in the caruncles and glands. Bovine trophoblast did not express integrin alpha V beta 3 at any stage of pregnancy. In ovine endometrium a different pattern of staining for integrin alpha V beta 3 was observed. Reactivity was not present in the luminal epithelium or trophoblast. There was strong staining of the deep glands and no reactivity in the superficial glands. Osteopontin distribution was similar for sheep and cattle. For both species, apical staining was present on the luminal epithelium and glands and on embryonic tissues. Conclusion In ruminants, integrin alpha V beta 3 and osteopontin do not co-localize at the fetal-maternal interface indicating that these proteins could not interact to facilitate embryo attachment as has been proposed in other species.

Histological spectrum of angiofibroma of soft tissue: histological and genetic analysis of 13 cases.

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Yamada, Yuichi; Yamamoto, Hidetaka; Kohashi, Kenichi; Ishii, Takeaki; Iura, Kunio; Maekawa, Akira; Bekki, Hirofumi; Otsuka, Hiroshi; Yamashita, Kyoko; Tanaka, Hiroyuki; Hiraki, Tsubasa; Mukai, Munenori; Shirakawa, Atsuko; Shinnou, Yoko; Jinno, Mari; Yanai, Hiroyuki; Taguchi, Kenichi; Maehara, Yoshihiko; Iwamoto, Yukihide; Oda, Yosinao

2016-09-01

Angiofibroma of soft tissue (AFST) is a rare soft tissue neoplasm characterized by a fibroblastic cytomorphology and a prominent vascular structure. AFSTs possess a novel fusion gene, i.e. NCOA2-AHRR/AHRR-NCOA2 or GTF2I-NCOA2, providing a useful approach to diagnosing AFST. Morphologically, AFSTs span a wide spectrum, making diagnosis a challenge. The aim of this study was to review AFST cases and to report previously unknown histological features, which we confirmed by genetic analysis. We reviewed 276 cases diagnosed as solitary fibrous tumours/haemangiopericytomas (232 cases), unclassified tumours of fibroblastic differentiation (36 cases), and recently diagnosed AFSTs (eight cases), and retrieved 13 cases compatible with AFST. Immunohistochemical staining was performed for these cases, all 13 of which were analysed by reverse transcription polymerase chain reaction and fluorescence in-situ hybridization. The histological findings were as follows: amianthoid fibres, extravasation of red blood cells, haemosiderin deposition, aggregates of foamy histiocytes, cystic change, necrosis, and haemorrhage. Immunohistochemically, the tumour cells were positive for epithelial membrane antigen (four of 13 cases), desmin (six of 13 cases), CD163 (13 of 13 cases), CD68 (seven of 13 cases), oestrogen receptor (13 of 13 cases), progesterone receptor (three of 13 cases), and STAT6 (one of 13 cases, weak nuclear staining), but they were negative for CD34, α-smooth muscle actin, muscle-specific actin, S100, pan-cytokeratin, MDM2, and CDK4. The AHRR-NCOA2 fusion gene was detected in eight cases, and NCOA2 gene rearrangement in nine cases. We revealed the previously unreported histological variation and immunohistochemical findings of AFST, and confirmed them by using genetic methods. The results suggested that AFST should be considered in the diagnosis of fibrous or fibrohistiocytic tumours with the above histological features. © 2016 John Wiley & Sons Ltd.

Immunohistochemical study on the expression of matrix metalloproteinase 2 and high-risk human papilloma virus in the malignant progression of papillomas

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Lee, Ho-Jin

2013-01-01

Objectives Papilloma frequently develops as a benign tumor of the head and neck area, but its potential for malignant transformation has yet to be studied. This study aims to provide basic information for papillomas using the immunohistochemical staining of matrix metalloproteinase 2 (MMP-2) and human papilloma virus (HPV) 16 and 18. Materials and Methods To evaluate the malignant transformation of papillomas, the selected tissue samples were serially diagnosed with pre-cancerous papilloma (with epithelial dysplasia, pseudo-epitheliomatous hyperplasia) or malignant lesion (squamous cell carcinoma, SCC) after the first diagnosis (squamous papilloma, inverted papilloma). The selected tissues were stained with an antibody to MMP-2 and HPV 16-E7, HPV 18-L1. A statistical analysis was performed according to each transformation step. Results The epithelial layer of papilloma and pre-cancerous papilloma lesions had a similar MMP-2 expression, but that of the malignant lesion had a significantly increased MMP-2 expression. HPV 16 and 18 infection rates were 28.6%, 33.3% and 63.6% in papillomas, pre-cancerous papilloma lesions, and SCC. Conclusions A relatively high MMP-2 expression and HPV 16 or 18 infection of papillomas may be associated with early events in the multistep processes of malignant transformation of papillomas. PMID:24471049

Automated quantification of proliferation with automated hot-spot selection in phosphohistone H3/MART1 dual-stained stage I/II melanoma.

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Nielsen, Patricia Switten; Riber-Hansen, Rikke; Schmidt, Henrik; Steiniche, Torben

2016-04-09

Staging of melanoma includes quantification of a proliferation index, i.e., presumed melanocytic mitoses of H&E stains are counted manually in hot spots. Yet, its reproducibility and prognostic impact increases by immunohistochemical dual staining for phosphohistone H3 (PHH3) and MART1, which also may enable fully automated quantification by image analysis. To ensure manageable workloads and repeatable measurements in modern pathology, the study aimed to present an automated quantification of proliferation with automated hot-spot selection in PHH3/MART1-stained melanomas. Formalin-fixed, paraffin-embedded tissue from 153 consecutive stage I/II melanoma patients was immunohistochemically dual-stained for PHH3 and MART1. Whole slide images were captured, and the number of PHH3/MART1-positive cells was manually and automatically counted in the global tumor area and in a manually and automatically selected hot spot, i.e., a fixed 1-mm(2) square. Bland-Altman plots and hypothesis tests compared manual and automated procedures, and the Cox proportional hazards model established their prognostic impact. The mean difference between manual and automated global counts was 2.9 cells/mm(2) (P = 0.0071) and 0.23 cells per hot spot (P = 0.96) for automated counts in manually and automatically selected hot spots. In 77 % of cases, manual and automated hot spots overlapped. Fully manual hot-spot counts yielded the highest prognostic performance with an adjusted hazard ratio of 5.5 (95 % CI, 1.3-24, P = 0.024) as opposed to 1.3 (95 % CI, 0.61-2.9, P = 0.47) for automated counts with automated hot spots. The automated index and automated hot-spot selection were highly correlated to their manual counterpart, but altogether their prognostic impact was noticeably reduced. Because correct recognition of only one PHH3/MART1-positive cell seems important, extremely high sensitivity and specificity of the algorithm is required for prognostic purposes. Thus, automated

Propidium iodide staining: a new application in fluorescence microscopy for analysis of cytoarchitecture in adult and developing rodent brain.

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Hezel, Marcus; Ebrahimi, Fahim; Koch, Marco; Dehghani, Faramarz

2012-10-01

Immunohistochemical visualization of antigens in specimen has evolved to an indispensable technique in biomedical research for investigations of cell morphology and pathology both in bright field and fluorescence microscopy. While there are couple of staining methods that reveal entire cytoarchitecture in bright field microscopy such as Nissl or hemalaun-eosin, there are still limitations in visualizations of cytoarchitecture in fluorescence microscopy. The present study reports a simple staining method that provides the required illustration of cell allocations and cellular composition in fluorescence microscopy in adult and in developing rodent central nervous system using the fluorophore propidium iodide (PI, 5μg/mL). PI is a well-accepted marker for degenerating cells when applied prior to fixation (pre-fixation PI staining). Here, PI was added to the sections after the fixation (post-fixation PI staining). This revised labeling procedure led to similar cytoarchitectural staining patterns in fluorescence microscopy as observed with hemalaun in bright field microscopy. This finding was proven in organotypic hippocampal slice cultures (OHSC) and brain sections obtained from different postnatal developmental stages. Excitotoxically lesioned OHSC subjected to pre-fixation PI staining merely showed brightly labeled condensed nuclei of degenerating neurons. In contrast, post-fixation PI staining additionally revealed extensive labeling of neuronal cell bodies and glial cells within the OHSC, thus allowing visualization of stratification of neuronal layers and cell morphology. Furthermore, post-fixation PI staining was combined with NeuN, calbindin, calretinin, glial fibrillary acidic protein or Griffonia simplicifolia isolectin B4 (IB(4)) in post natal (p1 and p9) and adult rats. In early post-natal brain sections almost all mentioned cellular markers led to an incomplete staining of the native cell organization and resulted in an inaccurate estimation of cell

Hierarchical patch-based co-registration of differently stained histopathology slides

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Yigitsoy, Mehmet; Schmidt, Günter

2017-03-01

Over the past decades, digital pathology has emerged as an alternative way of looking at the tissue at subcellular level. It enables multiplexed analysis of different cell types at micron level. Information about cell types can be extracted by staining sections of a tissue block using different markers. However, robust fusion of structural and functional information from different stains is necessary for reproducible multiplexed analysis. Such a fusion can be obtained via image co-registration by establishing spatial correspondences between tissue sections. Spatial correspondences can then be used to transfer various statistics about cell types between sections. However, the multi-modal nature of images and sparse distribution of interesting cell types pose several challenges for the registration of differently stained tissue sections. In this work, we propose a co-registration framework that efficiently addresses such challenges. We present a hierarchical patch-based registration of intensity normalized tissue sections. Preliminary experiments demonstrate the potential of the proposed technique for the fusion of multi-modal information from differently stained digital histopathology sections.

Immunohistochemical Correlation of Matrix Metalloproteinase-2 and Tissue Inhibitors of Metalloproteinase-2 in Tobacco Associated Epithelial Dysplasia

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Dipshikha Bajracharya

2014-01-01

Full Text Available Aim. To study the immunohistochemical expression of matrix metalloproteinase and tissue inhibitors of matrix metalloproteinase-2 in different histological grades of tobacco associated epithelial dysplasia and correlate the association between these proteases. Potentially malignant oral disorders (PMODs progressing to oral cancer are related to the severity of epithelial dysplasia. Methods. A retrospective immunohistochemical study was carried out on 30 clinically and histologically proven cases of leukoplakia with dysplasia and 10 cases of normal buccal mucosa using anti-MMP-2 and anti-TIMP-2 monoclonal antibodies. Results. Mann Whitney U test, for comparing the expression of both MMP-2 and TIMP-2 in normal mucosa with dysplasia, was highly significant (P<0.001. Kruskal-Wallis test to compare the median score of MMP-2 and TIMP-2 in different grades of dysplasia showed statistical significance (P<0.001, and a Spearman’s correlation between MMP-2 and TIMP-2 through different grades of dysplasia and cells observed showed positive correlation. Conclusion. Concomitant increase in the expression of both MMP-2 and TIMP-2 suggested that the activation of MMP-2 is dependent on TIMP-2 acting as a cofactor. Changes in TIMP-2 levels are considered important because they directly affect the level of MMP-2 activity.

[Comparative study of lymphoid follicles in mucosa of pharynx and mucosal associated lymphoid tissues in paranasal sinuses].

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Zhai, Weigang; Yao, Min; Chen, Jue

2013-08-01

To study the relationship between the lymphoid follicles in mucous membrane of pharynx and mucosal associated lymphoid tissues (MALT). Ten folliculi obtained from 10 patients of follicular pharyngitis and mucosa taken form 10 patients of paranasal sinusitis were fixed in neutral formalin and embedded in paraffin. Sections were prepared, stained by H. E and by immunohistochemical method staining with S-100,and observe by light microscopy. We observed the morphology of lymphoid follicles in mucous membrane of pharynx with MALT in mucosa of paranasal sinusitis as the contrast. Lymphoid follicles in mucosa of pharynx compared with MALT in the mucosa of paranasal sinuses, there was no mantle zone, no typical germinal center and no mucosal epithelium, immunological staining with S-100 was week. The lymphoid follicles in mucosa of pharynx does not belong to the MALT.

Oncocytic carcinoma of parotid gland: a case report with clinical, immunohistochemical and ultrastructural features

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Giordano Giovanna

2006-08-01

Full Text Available Abstract Background Oncocytic carcinoma is an extremely rare neoplasm of the salivary glands. We report a case of oncocytic carcinoma arising in a parotid gland in a 66-year-old female. Method An excisional biopsy of the parotid tumor was performed. The specimen was submitted for histology and after fixation in formalin solution and inclusion in paraffin, 3–5 μm sections were stained with hematoxylin and eosin for conventional evaluation and Periodic acid Schiff stain. Immunohistochemical studies were performed using antibodies against mitochondrial antigen, keratin, S-100, alpha-actin, vimentin, alpha-1-antichymotrypsin as well as an ultrastructural analysis was performed. Results Frozen sections revealed an infiltrative growth pattern and the diagnosis of a malignant epithelial lesion was made. Permanent sections stained with haematoxylin and eosin revealed a neoplasm that had replaced a wide area of the parotid gland and had invaded subcutaneous adipose tissue. Perineural invasion was evident, but vascular invasion was not found. Neoplastic elements were large, round or polyhedral cells and were arranged in solid sheets, islands and cords. The cytoplasm was abundant, eosinophilic and finely granular. The nuclei were large and located centrally or peripherally. The nucleoli were distinct and large. Periodic acid Schiff stain demonstrated a granular cytoplasm. Immunohistochemistry demonstrated mithochondrial antigen, keratin, and chymotrypsin immunoreactivity in the neoplastic cells. Ultrastructural analysis revealed numerous mitochondria packed into the cytoplasm of the neoplastic cells. Thus, the final diagnosis was that of oncocytic carcinoma of parotid gland. Conclusion This neoplasm shows clinical, microscopical, histological and ultrastructural features of oncocytic carcinoma and this must be considered in the differential diagnosis of other proliferations in the parotid gland with abundant granular cytoplasm and metastatic oncocytic

Clinicopathological and Immunohistochemical Characterisation of Gastric Schwannomas in 29 Cases

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Zheng, Lijun; Wu, Xiaojiang; Kreis, Martin E.; Yu, Zhen; Feng, Lijin; Chen, Chunqiu; Xu, Bin; Bu, Zhaode; Li, Ziyu; Ji, Jiafu

2014-01-01

Schwannomas are tumors arising from the nervous system that also occur infrequently in the gastrointestinal tract, most commonly in the stomach. This report characterizes 29 patients with benign or malignant gastric schwannomas. Surgical data and clinical follow-up information were available for 28 cases with a median postoperative duration of 57 months. Clinicopathological and immunohistochemical characteristics of benign and malignant schwannomas were analysed. Four cases (13.7%) were histologically diagnosed with malignant schwannoma. All tumors were positive for S-100 and CD56 proteins, displaying a diffuse staining pattern. Vimentin was expressed in 100% cases and all schwannomas were negative for smooth muscle actin, c-kit, and HMB-45. A significant difference was observed between the group of benign and malignant schwannomas as regards recurrences and metastasis after complete resection (P = 0.015). The survival time of patients with benign schwannomas was longer than the malignant group (P = 0.013), so gastric malignant schwannomas have a potential for recurrence and metastasis, with subsequently short survival. Complete resection with an attempt to remove all tumor tissue with negative margins is of paramount importance in the management of gastric schwannomas, particularly when they turn out to be malignant. PMID:24688535

Clinicopathological and Immunohistochemical Characterisation of Gastric Schwannomas in 29 Cases

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Lijun Zheng

2014-01-01

Full Text Available Schwannomas are tumors arising from the nervous system that also occur infrequently in the gastrointestinal tract, most commonly in the stomach. This report characterizes 29 patients with benign or malignant gastric schwannomas. Surgical data and clinical follow-up information were available for 28 cases with a median postoperative duration of 57 months. Clinicopathological and immunohistochemical characteristics of benign and malignant schwannomas were analysed. Four cases (13.7% were histologically diagnosed with malignant schwannoma. All tumors were positive for S-100 and CD56 proteins, displaying a diffuse staining pattern. Vimentin was expressed in 100% cases and all schwannomas were negative for smooth muscle actin, c-kit, and HMB-45. A significant difference was observed between the group of benign and malignant schwannomas as regards recurrences and metastasis after complete resection (P=0.015. The survival time of patients with benign schwannomas was longer than the malignant group (P=0.013, so gastric malignant schwannomas have a potential for recurrence and metastasis, with subsequently short survival. Complete resection with an attempt to remove all tumor tissue with negative margins is of paramount importance in the management of gastric schwannomas, particularly when they turn out to be malignant.

Antibody validation and scoring guidelines for ABCG2 immunohistochemical staining in formalin-fixed paraffin-embedded colon cancer tissue

DEFF Research Database (Denmark)

Cederbye, Camilla Natasha; Palshof, Jesper Andreas; Hansen, Tine Plato

2016-01-01

cancer (CRC), probably because of the use of different antibodies and scoring approaches. In this study, we systematically studied six commercially available anti-ABCG2 antibodies, using cell lines with up-regulation of ABCG2, and selected one antibody for validation in CRC tissue. Furthermore, we...... sections, especially when more than one core was used. In conclusion, here, we provide validated results to guide future studies on the associations between ABCG2 immunoreactivity in tumor cells and the benefits of chemotherapeutic treatment in patients with CRC...

Occurrence and significance of Mallory bodies in morbidly obese patients. An immunohistochemical study

DEFF Research Database (Denmark)

Gluud, C; Christoffersen, Pernille Yde; Andersen, T

1984-01-01

Liver biopsies from 61 consecutive patients with morbid obesity (less than 60% overweight) and from 48 patients with alcoholic liver disease were examined for the presence of Mallory bodies. For the detection both routine haematoxylin and eosin stained sections and sections exposed to an immunohi......Liver biopsies from 61 consecutive patients with morbid obesity (less than 60% overweight) and from 48 patients with alcoholic liver disease were examined for the presence of Mallory bodies. For the detection both routine haematoxylin and eosin stained sections and sections exposed...... to an immunohistochemical technique were employed. The latter uses an antiserum which recognizes antigenic determinants in Mallory bodies. Using haematoxylin and eosin staining. Mallory bodies were not detected in any of the biopsies from the obese patients, but found to be present in 63% of the patients with alcoholic...

One-Step Preservation of Phosphoproteins and Tissue Morphology at Room Temperature for Diagnostic and Research Specimens

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Mueller, Claudius; Edmiston, Kirsten H.; Carpenter, Calvin; Gaffney, Eoin; Ryan, Ciara; Ward, Ronan; White, Susan; Memeo, Lorenzo; Colarossi, Cristina; Petricoin, Emanuel F.; Liotta, Lance A.; Espina, Virginia

2011-01-01

Background There is an urgent need to measure phosphorylated cell signaling proteins in cancer tissue for the individualization of molecular targeted kinase inhibitor therapy. However, phosphoproteins fluctuate rapidly following tissue procurement. Snap-freezing preserves phosphoproteins, but is unavailable in most clinics and compromises diagnostic morphology. Formalin fixation preserves tissue histomorphology, but penetrates tissue slowly, and is unsuitable for stabilizing phosphoproteins. We originated and evaluated a novel one-step biomarker and histology preservative (BHP) chemistry that stabilizes signaling protein phosphorylation and retains formalin-like tissue histomorphology with equivalent immunohistochemistry in a single paraffin block. Results Total protein yield extracted from BHP-fixed, routine paraffin-embedded mouse liver was 100% compared to snap-frozen tissue. The abundance of 14 phosphorylated proteins was found to be stable over extended fixation times in BHP fixed paraffin embedded human colon mucosa. Compared to matched snap-frozen tissue, 8 phosphoproteins were equally preserved in mouse liver, while AMPKβ1 Ser108 was slightly elevated after BHP fixation. More than 25 tissues from mouse, cat and human specimens were evaluated for preservation of histomorphology. Selected tissues were evaluated in a multi-site, independent pathology review. Tissue fixed with BHP showed equivalent preservation of cytoplasmic and membrane cytomorphology, with significantly better nuclear chromatin preservation by BHP compared to formalin. Immunohistochemical staining of 13 non-phosphorylated proteins, including estrogen receptor alpha, progesterone receptor, Ki-67 and Her2, was equal to or stronger in BHP compared to formalin. BHP demonstrated significantly improved immunohistochemical detection of phosphorylated proteins ERK Thr202/Tyr204, GSK3-α/β Ser21/Ser9, p38-MAPK Thr180/Tyr182, eIF4G Ser1108 and Acetyl-CoA Carboxylase Ser79. Conclusion In a single

One-step preservation of phosphoproteins and tissue morphology at room temperature for diagnostic and research specimens.

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Claudius Mueller

Full Text Available BACKGROUND: There is an urgent need to measure phosphorylated cell signaling proteins in cancer tissue for the individualization of molecular targeted kinase inhibitor therapy. However, phosphoproteins fluctuate rapidly following tissue procurement. Snap-freezing preserves phosphoproteins, but is unavailable in most clinics and compromises diagnostic morphology. Formalin fixation preserves tissue histomorphology, but penetrates tissue slowly, and is unsuitable for stabilizing phosphoproteins. We originated and evaluated a novel one-step biomarker and histology preservative (BHP chemistry that stabilizes signaling protein phosphorylation and retains formalin-like tissue histomorphology with equivalent immunohistochemistry in a single paraffin block. RESULTS: Total protein yield extracted from BHP-fixed, routine paraffin-embedded mouse liver was 100% compared to snap-frozen tissue. The abundance of 14 phosphorylated proteins was found to be stable over extended fixation times in BHP fixed paraffin embedded human colon mucosa. Compared to matched snap-frozen tissue, 8 phosphoproteins were equally preserved in mouse liver, while AMPKβ1 Ser108 was slightly elevated after BHP fixation. More than 25 tissues from mouse, cat and human specimens were evaluated for preservation of histomorphology. Selected tissues were evaluated in a multi-site, independent pathology review. Tissue fixed with BHP showed equivalent preservation of cytoplasmic and membrane cytomorphology, with significantly better nuclear chromatin preservation by BHP compared to formalin. Immunohistochemical staining of 13 non-phosphorylated proteins, including estrogen receptor alpha, progesterone receptor, Ki-67 and Her2, was equal to or stronger in BHP compared to formalin. BHP demonstrated significantly improved immunohistochemical detection of phosphorylated proteins ERK Thr202/Tyr204, GSK3-α/β Ser21/Ser9, p38-MAPK Thr180/Tyr182, eIF4G Ser1108 and Acetyl-CoA Carboxylase Ser79

Staining plastic blocks with triiodide to image cells and soft tissues in backscattered electron SEM of skeletal and dental tissues

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A Boyde

2012-07-01

Full Text Available Backscattered electron scanning electron microscopy (BSE SEM is an invaluable method for studying the histology of the hard, mineralised components of poly-methyl methacrylate (PMMA or other resin embedded skeletal and dental tissues. Intact tissues are studied in micro-milled or polished block faces with an electron-optical section thickness of the order of a half to one micron and with the area of the section as big as a whole – large or small – bone organ. However, BSE SEM does not give information concerning the distribution of uncalcified, ‘soft’, cellular and extracellular matrix components. This can be obtained by confocal microscopy of the same block and the two sorts of images merged but the blocks have to be studied in two microscope systems. The present work shows a new, simple and economic approach to visualising both components by using the triiodide ion in Lugol's iodine solution to stain the block surface prior to the application of any conductive coating – and the latter can be omitted if charging is suppressed by use of poor vacuum conditions in the SEM sample chamber. The method permits the use of archival tissue, and it will be valuable in studies of both normal growth and development and pathological changes in bones and joints, including osteoporosis and osteoarthritis, and tissue adaptation to implants.

Demonstration of lipofuscin and Nissl bodies in crystal violet stained sections using a fluorescence technique or pyronin Y stain.

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Terr, L I

1986-09-01

This paper presents two simple, reliable methods for identification of lipofuscin and Nissl bodies in the same section. One method shows that lipofuscin stained with crystal violet retains its ability to fluoresce and can be observed under the fluorescence microscope after the stain has faded. Fading is accompanied by a gradual increase in the intensity of the fluorescence and is complete in about 5 min. Exciting illumination from this part of the spectrum also substantially fades staining of other autofluorescing tissue elements, such as lipids. Nonfluorescing structures, such as Nissl bodies, remain stained. By changing from transillumination with tungsten light to epifluorescent illumination and vice versa, both types of structures--Nissl bodies and lipofuscin--can be identified in the same section. The second technique uses pyronin Y for staining Nissl bodies in preparations previously stained with crystal violet. Nissl bodies are stained pink but lipofuscin remains violet. Lipofuscin in these sections also remains autofluorescent after the crystal violet stain has faded under violet or near-UV light.

Immunohistochemical Analysis of Human Vallate Taste Buds.

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Tizzano, Marco; Grigereit, Laura; Shultz, Nicole; Clary, Matthew S; Finger, Thomas E

2015-11-01

The morphology of the vallate papillae from postmortem human samples was investigated with immunohistochemistry. Microscopically, taste buds were present along the inner wall of the papilla, and in some cases in the outer wall as well. The typical taste cell markers PLCβ2, GNAT3 (gustducin) and the T1R3 receptor stain elongated cells in human taste buds consistent with the Type II cells in rodents. In the human tissue, taste bud cells that stain with Type II cell markers, PLCβ2 and GNAT3, also stain with villin antibody. Two typical immunochemical markers for Type III taste cells in rodents, PGP9.5 and SNAP25, fail to stain any taste bud cells in the human postmortem tissue, although these antibodies do stain numerous nerve fibers throughout the specimen. Car4, another Type III cell marker, reacted with only a few taste cells in our samples. Finally, human vallate papillae have a general network of innervation similar to rodents and antibodies directed against SNAP25, PGP9.5, acetylated tubulin and P2X3 all stain free perigemmal nerve endings as well as intragemmal taste fibers. We conclude that with the exception of certain molecular features of Type III cells, human vallate papillae share the structural, morphological, and molecular features observed in rodents. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Immunohistochemical Expression of P53 Protein in Cutaneous Basal Cell Carcinoma: A Clinicopathological Study of 66 Cases

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Vladim and iacute;r Barto and scaron;

2016-12-01

Full Text Available Objective: Nuclear expression of p53 protein is associated with a biological behavior in a variety of human malignancies. In cutaneous basal cell carcinoma (BCC, however, many studies have provided conflicting results in this regard. We aimed to determine whether there is relationship between p53 expression and different histologic subtypes of BCC, and whether it may indicate tumor aggressiveness. Materials and Methods: Biopsy samples from 66 cutaneous BCCs from 57 patients were collected. P53 expression was demonstrated by immunohistochemical staining using the anti-p53 antibody. Among them, 52 cases were also evaluated for Ki-67 antigen. Results: Immunoreactivity of p53 protein varied in the range of 0 to 100% of total tumor tissue (mean value 46.0%. The expression exceeding 5% of cancer tissue (positive staining was found in 54 BCCs (81.8%. Within this group, there were 25 cases (37.9% with low and 29 cases (43.9% with high expression. In superficial, superficial-nodular, nodular, nodular-infiltrative and infiltrative BCCs, p53 protein positivity was found in 100% (8/8, 80% (8/10, 70.4% (19/27, 88.2% (15/17 and 100% (4/4, respectively. We did not reveal a significant correlation between the extent of p53 protein expression and BCC subtypes except for nodular BCC, in which a number of negative cases (8/27, 29.6% were just above the threshold of statistical significance (P = 0.04. After merging cancers into non-aggressive and aggressive growth phenotype, no association with expression of p53 protein was found. There was no relationship between p53 protein expression and topographical sites after they have been gathered into sun-exposed and sun-protected locations. We did not observe any association between expression of p53 protein and Ki-67 antigen. Conclusion: In cutaneous BCC, the expression of p53 protein does not seem to reflect a biological behavior and tumor aggressiveness. Therefore, in a routine dermatopathological practice

Nucleic Acid Amplification Testing and Sequencing Combined with Acid-Fast Staining in Needle Biopsy Lung Tissues for the Diagnosis of Smear-Negative Pulmonary Tuberculosis.

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Faming Jiang

Full Text Available Smear-negative pulmonary tuberculosis (PTB is common and difficult to diagnose. In this study, we investigated the diagnostic value of nucleic acid amplification testing and sequencing combined with acid-fast bacteria (AFB staining of needle biopsy lung tissues for patients with suspected smear-negative PTB.Patients with suspected smear-negative PTB who underwent percutaneous transthoracic needle biopsy between May 1, 2012, and June 30, 2015, were enrolled in this retrospective study. Patients with AFB in sputum smears were excluded. All lung biopsy specimens were fixed in formalin, embedded in paraffin, and subjected to acid-fast staining and tuberculous polymerase chain reaction (TB-PCR. For patients with positive AFB and negative TB-PCR results in lung tissues, probe assays and 16S rRNA sequencing were used for identification of nontuberculous mycobacteria (NTM. The sensitivity, specificity, positive predictive value (PPV, negative predictive value (NPV, and diagnostic accuracy of PCR and AFB staining were calculated separately and in combination.Among the 220 eligible patients, 133 were diagnosed with TB (men/women: 76/57; age range: 17-80 years, confirmed TB: 9, probable TB: 124. Forty-eight patients who were diagnosed with other specific diseases were assigned as negative controls, and 39 patients with indeterminate final diagnosis were excluded from statistical analysis. The sensitivity, specificity, PPV, NPV, and accuracy of histological AFB (HAFB for the diagnosis of smear-negative were 61.7% (82/133, 100% (48/48, 100% (82/82, 48.5% (48/181, and 71.8% (130/181, respectively. The sensitivity, specificity, PPV, and NPV of histological PCR were 89.5% (119/133, 95.8% (46/48, 98.3% (119/121, and 76.7% (46/60, respectively, demonstrating that histological PCR had significantly higher accuracy (91.2% [165/181] than histological acid-fast staining (71.8% [130/181], P < 0.001. Parallel testing of histological AFB staining and PCR showed the

Immunohistochemical Analysis of Vimentin in Oral Sub mucous Fibrosis

International Nuclear Information System (INIS)

Nayak, M. T.; Singh, A.; Desai, R. S.; Vanaki, S. S.

2013-01-01

Oral sub mucous fibrosis (OSF), a precancerous condition, is characterized by abnormal accumulation of collagen fibers in oral sub mucosa. Vimentin is a Class 2 intermediate filament (IF) and primarily expressed in cells of mesenchymal origin. Vimentin is also found to be involved in cell growth, cell cycling, and tumour differentiation. Objective. The purpose of the study was to compare the expression of vimentin in various histological grades of OSF. Materials and Methods. To assess the immunohistochemical expression of vimentin in 20 mild cases of OSF, 20 severe cases of OSF, and ten cases of normal oral buccal mucosa. Results. The overall staining intensity of vimentin significantly increased statistically (P<0.01) in OSF cases over normal control. A significant increase in the staining intensity of vimentin was also noted in the fibroblasts of severe cases of OSF (P=0.03). Conclusion. Considering the marked vimentin expression in the present study, future studies should include cytoskeleton IF and other filaments in the fibroblasts of OSF.

Immunohistochemical assessment of oestrogen and progesterone receptors

DEFF Research Database (Denmark)

Grabau, D A; Thorpe, S M; Knoop, A

2000-01-01

Two different methods to determine steroid receptors were analysed with respect to their ability to estimate prognosis in primary breast cancer patients. The immunohistochemical assay (IHA) was compared with the dextran-coated charcoal (DCC) method of receptor determination. A random sample of 28...... distinction between benign and malignant tissue is possible using the IHAmethod. Thus, IHAresults appear to be more clinically relevant....

Cell-surface glycoproteins of human sarcomas: differential expression in normal and malignant tissues and cultured cells

International Nuclear Information System (INIS)

Rettig, W.F.; Garin-Chesa, P.; Beresford, H.R.; Oettgen, H.F.; Melamed, M.R.; Old, L.J.

1988-01-01

Normal differentiation and malignant transformation of human cells are characterized by specific changes in surface antigen phenotype. In the present study, the authors have defined six cell-surface antigens of human sarcomas and normal mesenchymal cells, by using mixed hemadsorption assays and immunochemical methods for the analysis of cultured cells and immunohistochemical staining for the analysis of normal tissues and > 200 tumor specimens. Differential patterns of F19, F24, G171, G253, S5, and Thy-1 antigen expression were found to characterize (i) subsets of cultured sarcoma cell lines, (ii) cultured fibroblasts derived from various organs, (iii) normal resting and activated mesenchymal tissues, and (iv) sarcoma and nonmesenchymal tumor tissues. These results provide a basic surface antigenic map for cultured mesenchymal cells and mesenchymal tissues and permit the classification of human sarcomas according to their antigenic phenotypes

Correlation of Ki-67, p53, and Adnab-9 immunohistochemical staining and ploidy with clinical and histopathologic features of severely dysplastic colorectal adenomas.

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Sheikh, Rafiq A; Min, Byung Hee; Yasmeen, Shagufta; Teplitz, Raymond; Tesluk, Henry; Ruebner, Boris Henry; Tobi, Martin; Hatfield, James; Fligiel, Suzanne; Lawson, Michael J

2003-01-01

Variations of Ki-67, p53, and Adnab-9 monoclonal antibody reactions in colonic adenomas may be associated with colonic cancer risk. We studied the predictive value of these markers for adverse behavior in severely dysplastic colorectal adenomas, such as an associated carcinoma, multiplicity of adenomas, and subsequent development of adenomas. For this purpose we compared theclinical, gross, and histologic characteristics of highly dysplastic index polyps in 42 patients with Ki 67, p53, and Adnab-9 immunostaining and other molecular markers. Polyps were removed endoscopically, and severely dysplastic polyps were stained immunohistochemically with Ki-67, Adnab-9, and p53 protein by the avidin biotin conjugate (ABC) technique. Quantitative DNA (QDNA) was analyzed by computer-assisted image analysis. Ki-67 immunohistochemistry showed reversal of normal distribution of nuclear staining from the normal basal position to the upper third of the colonic crypts. This abnormality of immunostaining in dysplastic adenomas was the earliest detected by the panel we used. A statistically significant correlation was seen between invasiveness of carcinoma in the index polyp and polyp size (P = 0.003), sessile morphology (P = 0.037), and villous or tubulovillous histology (P = 0.019). In the index adenoma, p53 positivity was correlated with multiplicity at initial examination (P = 0.053), villous histology (P = 0.053), invasiveness of carcinoma (P < 0.003), and recurrence of colorectal adenomas (P = 0.025). Although p53 positivity and aneuploidy were correlated with invasiveness of carcinoma in the index polyp (P = 0.025), Adnab-9 positivity was not. However, Adnab-9 positivity in the index polyp was associated with multiplicity of adenomas (P = 0.04) as well as recurrence of adenomas (P < 0.024). In conclusion, in addition to the morphologic and histologic markers already known, Ki-67, Adnab-9 antibody, and p53 protein may be prognostic indicators useful in follow-up of patients

Characterization of cells from pannus-like tissue over articular cartilage of advanced osteoarthritis.

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Yuan, G-H; Tanaka, M; Masuko-Hongo, K; Shibakawa, A; Kato, T; Nishioka, K; Nakamura, H

2004-01-01

To identify the characteristics of cells isolated from pannus-like soft tissue on osteoarthritic cartilage (OA pannus cells), and to evaluate the role of this tissue in osteoarthritis (OA). OA pannus cells were isolated from pannus-like tissues in five joints obtained during arthroplasty. The phenotypic features of the isolated cells were characterized by safranin-O staining and immunohistochemical studies. Expression of MMP-1, MMP-3 and MMP-13 was also assessed using reverse transcriptase-polymerase chain reactions (RT-PCR), enzyme-linked immunosorbent assay (ELISA) and immunocytochemistry. Foci and plaque formation of pannus-like tissue over cartilage surface were found in 15 of 21 (71.4%) OA joints macroscopically, and among them, only five samples had enough tissue to be isolated. OA pannus cells were positive for type I collagen and vimentin, besides they also expressed type II collagen and aggrecan mRNA. Spontaneous expression of MMP-1, MMP-3 and MMP-13 was detected in OA pannus cells. Similar or higher levels of MMPs were detected in the supernatant of cultured OA pannus cells compared to OA chondrocytes, and among these MMP-3 levels were relatively higher in OA pannus cells. Immunohistochemically, MMP-3 positive cells located preferentially in pannus-like tissue on the border of original hyaline cartilage. Our results showed that OA pannus cells shared the property of mesenchymal cells and chondrocytes; however, their origin seemed different from chondrocytes or synoviocytes. The spontaneous expression of MMPs suggests that they are involved in the articular degradation in OA.

Vascularization after treatment of gingival recession defects with platelet-rich fibrin or connective tissue graft.

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Eren, Gülnihal; Kantarcı, Alpdoğan; Sculean, Anton; Atilla, Gül

2016-11-01

The aim of this study was to evaluate histologically the following treatment of bilateral localized gingival recessions with coronally advanced flap (CAF) combined with platelet-rich fibrin (PRF) or subepithelial connective tissue graft (SCTG). Tissue samples were harvested from 14 subjects either 1 or 6 months after the surgeries. The 2-mm punch biopsies were obtained from the mid-portion of the grafted sites. Neutral buffered formalin fixed, paraffin-embedded 5-μm thick tissue sections were stained with hematoxylin eosin and Masson's trichrome in order to analyze the collagen framework, epithelium thickness and rete-peg length. Multiple sequential sections were cut from paraffin-embedded blocks of tissue and immunohistochemically prepared for detection of vascular endothelial growth factor, CD31 and CD34, for the assessment of vascularization. Rete peg formation was significantly increased in the sites treated with PRF compared to the SCTG group after 6 months (p < 0.05). On the contrary, the number of vessels was increased in the SCTG group compared to the PRF group after 6 months (p < 0.05). No statistically significant differences were observed in the collagen density. Staining intensity of CD31 increased in submucosal area of PRF group than SCTG group after 1 month. Higher staining intensity of CD34 was observed in the submucosal area of PRF group compared with SCTG group after 6 months. The results of the present study suggest that in histological evaluation because of its biological compounds, PRF results earlier vessel formation and tissue maturation compared to connective tissue graft. PRF regulated the vascular response associated with an earlier wound healing.

Expression of human telomerase reverse transcriptase protein in oral epithelial dysplasia and oral squamous cell carcinoma: An immunohistochemical study

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Raghunandan, Bangalore Nagarajachar; Sanjai, Karpagaselvi; Kumaraswamy, Jayalakshmi; Papaiah, Lokesh; Pandey, Bhavna; Jyothi, Bellur MadhavaRao

2016-01-01

Background: Telomerase is an RNA-dependent DNA polymerase that synthesizes TTAGGG telomeric DNA sequences and almost universally provides the molecular basis for unlimited proliferative potential. The telomeres become shorter with each cycle of replication and reach a critical limit; most cells die or enter stage of replicative senescence. Telomere length maintenance by telomerase is required for all the cells that exhibit limitless replicative potential. It has been postulated that reactivation of telomerase expression is necessary for the continuous proliferation of neoplastic cells to attain immortality. Use of immunohistochemistry (IHC) is a useful, reliable method of localizing the human telomerase reverse transcriptase (hTERT) protein in tissue sections which permits cellular localization. Although there exists a lot of information on telomerase in oral cancer, little is known about their expression in oral epithelial dysplasia and their progression to oral squamous cell carcinoma (OSCC) compared to normal oral mucosa. This study addresses this lacuna. Aims: To compare the expression of hTERT protein in oral epithelial dysplasia and OSCC with normal oral mucosa by Immunohistochemical method. Subjects and Methods: In this preliminary study, IHC was used to detect the expression of hTERT protein in OSCC (n = 20), oral epithelial dysplasia (n = 21) and normal oral mucosa (n = 10). The tissue localization of immunostain, cellular localization of immunostain, nature of stain, intensity of stain, percentage of cells stained with hTERT protein were studied. A total number of 100 cells were counted in each slide. Statistical Analysis: All the data were analyzed using SPSS software version 16.0. The tissue localization, cellular localization of cytoplasmic/nuclear/both of hTERT stain, staining intensity was compared across the groups using Pearson's Chi-square test. The mean percentage of cells stained for oral epithelial dysplasia, OSCC and normal oral mucosa were

Staining Methods for Normal and Regenerative Myelin in the Nervous System.

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Carriel, Víctor; Campos, Antonio; Alaminos, Miguel; Raimondo, Stefania; Geuna, Stefano

2017-01-01

Histochemical techniques enable the specific identification of myelin by light microscopy. Here we describe three histochemical methods for the staining of myelin suitable for formalin-fixed and paraffin-embedded materials. The first method is conventional luxol fast blue (LFB) method which stains myelin in blue and Nissl bodies and mast cells in purple. The second method is a LBF-based method called MCOLL, which specifically stains the myelin as well the collagen fibers and cells, giving an integrated overview of the histology and myelin content of the tissue. Finally, we describe the osmium tetroxide method, which consist in the osmication of previously fixed tissues. Osmication is performed prior the embedding of tissues in paraffin giving a permanent positive reaction for myelin as well as other lipids present in the tissue.

Sarcoglycans in the normal and pathological breast tissue of humans: an immunohistochemical and molecular study.

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Arco, Alba; Favaloro, Angelo; Gioffrè, Mara; Santoro, Giuseppe; Speciale, Francesco; Vermiglio, Giovanna; Cutroneo, Giuseppina

2012-01-01

The sarcoglycan complex, consisting of α-, β-, γ-, δ- and ε-sarcoglycans, is a multimember transmembrane system providing a mechanosignaling connection from the cytoskeleton to the extracellular matrix. Whereas the expression of α- and γ-sarcoglycan is restricted to striated muscle, other sarcoglycans are widely expressed. Although many studies have investigated sarcoglycans in all muscle types, insufficient data are available on the distribution of the sarcoglycan complex in nonmuscle tissue. On this basis, we used immunohistochemical and RT-PCR techniques to study preliminarily the sarcoglycans in normal glandular breast tissue (which has never been studied in the literature on these proteins) to verify the effective wider distribution of this complex. Moreover, to understand the role of sarcoglycans, we also tested samples obtained from patients affected by fibrocystic mastopathy and breast fibroadenoma. Our data showed, for the first time, that all sarcoglycans are always detectable in all normal samples both in epithelial and myoepithelial cells; in pathological breast tissue, all sarcoglycans appeared severely reduced. These data demonstrated that all sarcoglycans, not only β-, δ-, and ε-sarcoglycans, have a wider distribution, implying a new unknown role for these proteins. Moreover, in breast diseases, sarcoglycans containing cadherin domain homologs could provoke a loss of strong adhesion between epithelial cells, permitting and facilitating the degeneration of these benign breast tumors into malignant tumors. Consequently, sarcoglycans could play an important and intriguing role in many breast diseases and in particular in tumor progression from benign to malignant. Copyright © 2011 S. Karger AG, Basel.

Improved method for combination of immunocytochemistry and Nissl staining.

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Kádár, Andrea; Wittmann, Gábor; Liposits, Zsolt; Fekete, Csaba

2009-10-30

Nissl staining is a widely used method to study morphology and pathology of neural tissue. After standard immunocytochemistry, the Nissl staining labels only the nucleus of neurons and the characteristic staining of the neuronal perikarya is absent or very weak. We hypothesized that the RNA degradation during the immunocytochemical treatment results in the loss of cytoplasmic staining with Nissl-dyes. To test this hypothesis, we used RNAse-free conditions for all steps of immunostaining. To further prevent the RNA-degradation by RNAse contaminations, the RNAse inhibitor heparin was added to all antibody-containing solutions. The efficiency of Nissl staining after standard and RNAse-free double-labeling immunocytochemistry was compared using antibodies against c-Fos and neuropeptide Y (NPY) on tissues of rats refed after 3 days of fasting. After standard immunocytochemistry, the Nissl-staining labeled the nuclei of neurons and only very faintly the cytoplasm of these cells. The RNAse-free treatment did not alter the distribution of immunoreaction signal, but preserved the staining of neuronal perikarya by the Nissl-dyes. In conclusion, the RNAse-free conditions during immunocytochemistry allow the labeling of neuronal perikarya by Nissl-dyes. The described method facilitates the mapping of immunocytochemical signals and makes possible the light microscopic examination of the innervation of neurons identified by their nuclear protein content.

Epithelium percentage estimation facilitates epithelial quantitative protein measurement in tissue specimens.

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Chen, Jing; Toghi Eshghi, Shadi; Bova, George Steven; Li, Qing Kay; Li, Xingde; Zhang, Hui

2013-12-01

The rapid advancement of high-throughput tools for quantitative measurement of proteins has demonstrated the potential for the identification of proteins associated with cancer. However, the quantitative results on cancer tissue specimens are usually confounded by tissue heterogeneity, e.g. regions with cancer usually have significantly higher epithelium content yet lower stromal content. It is therefore necessary to develop a tool to facilitate the interpretation of the results of protein measurements in tissue specimens. Epithelial cell adhesion molecule (EpCAM) and cathepsin L (CTSL) are two epithelial proteins whose expressions in normal and tumorous prostate tissues were confirmed by measuring staining intensity with immunohistochemical staining (IHC). The expressions of these proteins were measured by ELISA in protein extracts from OCT embedded frozen prostate tissues. To eliminate the influence of tissue heterogeneity on epithelial protein quantification measured by ELISA, a color-based segmentation method was developed in-house for estimation of epithelium content using H&E histology slides from the same prostate tissues and the estimated epithelium percentage was used to normalize the ELISA results. The epithelium contents of the same slides were also estimated by a pathologist and used to normalize the ELISA results. The computer based results were compared with the pathologist's reading. We found that both EpCAM and CTSL levels, measured by ELISA assays itself, were greatly affected by epithelium content in the tissue specimens. Without adjusting for epithelium percentage, both EpCAM and CTSL levels appeared significantly higher in tumor tissues than normal tissues with a p value less than 0.001. However, after normalization by the epithelium percentage, ELISA measurements of both EpCAM and CTSL were in agreement with IHC staining results, showing a significant increase only in EpCAM with no difference in CTSL expression in cancer tissues. These results

The Immunohistochemical Analysis of SOCS3 Protein Identifies a Subgroup of Prostatic Cancer Biopsies With Aggressive Behavior.

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Pierconti, Francesco; Martini, Maurizio; Cenci, Tonia; Larocca, Luigi M

Recently, we demonstrated that hypermethylation of SOCS3 determines a significant reduction of its mRNA and protein expression and identifies a subgroup of prostate cancer with aggressive behavior. In this paper, our objective was to investigate whether the immunohistochemical expression of the SOCS3 protein could represent an alternative method to molecular analysis for the individualization of aggressive prostate carcinoma. We analyzed the SOCS3 immunohistochemical expression in 65 patients undergoing biopsies at the Institute of Urology of our hospital between September 2011 and October 2011 (median age, 66.4 y; range, 50 to 73 y), and in 35 cases, a subset of 65 cases originally used for the immunohistochemical study, we studied the methylation status of the SOCS3 promoter. We found that the percentage of cases with SOCS3 negativity (-) or with SOCS3 weak staining in <50% of the neoplastic glands (+/-) correlated to the worst prognosis in terms of the Gleason score (P=0.0001; Fisher's exact test), the pT stage (P=0.012; Fisher's exact test), and progression-free survival (P=0.0334; hazard ratio, 0.34; and 95% confidence interval, from 0.1261 to 0.9188). Moreover, some cases with an SOCS3 unmethylated pattern showed SOCS3-negative immunostaining (-) or SOCS3-negative glands with weak cytoplasmatic staining in <50% of the neoplastic glands (+/-). Our data suggest that in prostatic cancer biopsies, the immunohistochemical analysis of SOCS3 protein expression may provide a method that is less expensive and easier to apply than SOCS3 methylation analysis for the distinction of a subgroup of prostate cancer with a more aggressive behavior.

Immunohistochemical features of multifocal melanoacanthoma in the hard palate: a case report

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das Chagas e Silva de Carvalho Luis Felipe

2013-01-01

Full Text Available Abstract Background Melanoacanthoma (MA has been described in the oral mucosa as a solitary lesion or, occasionally, as multiple lesions. MA mainly affects dark skinned patients and grows rapidly, showing a plane or slightly raised appearance and a brown to black color. The differential diagnosis includes oral nevi, amalgam tattoos, and melanomas. We report here the case of a 58-year-old black woman who presented multiple pigmented lesions on the hard palate. Case presentation Based on the differential diagnosis of melanoma, a punch biopsy (4 mm in diameter was performed. The material was fixed in 10% formalin, embedded in paraffin, and stained with hematoxylin-eosin or submitted to immunohistochemical analysis. Immunohistochemistry using antibodies against protein S-100, melan-A, HMB-45, MCM-2, MCM-5, Ki-67 and geminin was performed. Immunohistochemical analysis revealed strong cytoplasmic immunoreactivity of dendritic melanocytes for proteinS-100, HMB-45 and melan-A.Positive staining for proliferative markers (MCM-2, MCM-5, Ki-67 was only observed in basal and suprabasal epithelial cells, confirming the reactive etiology of the lesion. The diagnosis was oral Melanoacanthoma (MA. Conclusion The patient has been followed up for 30 months and shows no clinical alterations. MA should be included in the differential diagnosis of pigmented lesions of the oral cavity.

Utility of Modified Ultrafast Papanicolaou Stain in Cytological Diagnosis.

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Sinkar, Prachi; Arakeri, Surekha Ulhas

2017-03-01

Need for minimal turnaround time for assessing Fine Needle Aspiration Cytology (FNAC) has encouraged innovations in staining techniques that require lesser staining time with unequivocal cell morphology. The standard protocol for conventional Papanicolaou (PAP) stain requires about 40 minutes. To overcome this, Ultrafast Papanicolaou (UFP) stain was introduced which reduces staining time to 90 seconds and also enhances the quality. However, reagents required for this were not easily available hence, Modified Ultrafast Papanicolaou (MUFP) stain was introduced subsequently. To assess the efficacy of MUFP staining by comparing the quality of MUFP stain with conventional PAP stain. FNAC procedure was performed by using 10 ml disposable syringe and 22-23 G needle. Total 131 FNAC cases were studied which were lymph node (30), thyroid (38), breast (22), skin and soft tissue (24), salivary gland (11) and visceral organs (6). Two smears were prepared and stained by MUFP and conventional PAP stain. Scores were given on four parameters: background of smears, overall staining pattern, cell morphology and nuclear staining. Quality Index (QI) was calculated from ratio of total score achieved to maximum score possible. Statistical analysis using chi square test was applied to each of the four parameters before obtaining the QI in both stains. Students t-test was applied to evaluate the efficacy of MUFP in comparison with conventional PAP stain. The QI of MUFP for thyroid, breast, lymph node, skin and soft tissue, salivary gland and visceral organs was 0.89, 0.85, 0.89, 0.83, 0.92, and 0.78 respectively. Compared to conventional PAP stain QI of MUFP smears was better in all except visceral organ cases and was statistically significant. MUFP showed clear red blood cell background, transparent cytoplasm and crisp nuclear features. MUFP is fast, reliable and can be done with locally available reagents with unequivocal morphology which is the need of the hour for a cytopathology set-up.

TEM validation of immunohistochemical staining prior to assessment of tumour angiogenesis by computerised image analysis

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Killingsworth, M.C.

2002-01-01

Full text: Counts of microvessel density (MVD) within solid tumours have been shown to be an independent predictor of outcome with higher counts generally associated with a worse prognosis. These assessments are commonly performed on immunoperoxidase stained (IPX) sections with antibodies to CD34, CD31 and Factor VIII-related antigen routinely used as vascular markers. Tumour vascular density is thought to reflect the demand the growing neoplasm is placing on its feeding blood supply. Vascular density also appears to be associated with spread of invasive cells to distant sites. The present study of tumour angiogenesis in prostate cancer specimens aims to assess new vessel growth in addition to MVD counts. The hypothesis being that an assessment which takes into account vascular migration and proliferation as well as the number of patent vessels present may have improved predictive power over assessments based on MVD counts alone. We are employing anti-CD34 stained IPX sections which are digitally photographed and assessed by a computerised image analysis system. Our aim is to develop parameters whereby tumour angiogenesis may be assessed at the light microscopic level and then correlated with existing histological methods of tumour assessment such as Gleason grading. In order to use IPX stained sections for angiogenic assessment validation and understanding of the anti-CD34 immunostaining pattern was necessary. This involved the following steps: i) Morphological assessment of angiogenic changes present in tumour blood vessels. Morphological changes in endothelial cells and pericytes indicative of angiogenic activation are generally below the level of resolution available with light microscopy. TEM examination revealed endothelial cell budding, pericyte retraction, basement membrane duplication and endothelial sprout formation in capillaries and venules surrounding tumour glands. This information assisted with the development of parameters by which IPX sections

Fluorescent Staining of Tea Pathogenic Fungi in Tea Leaves Using Fluorescein-labeled Lectin

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Yamada, Kengo; Yoshida, Katsuyuki; Sonoda, Ryoichi

Fluorochrome-labeled lectin, fluorescein conjugated wheat germ agglutinin (F-WGA) was applied to stain tea pathogenic fungi in tea leaf tissue. Infected leaves were fixed and decolorized with a mixture of ethanol and acetic acid, and cleared with 10% KOH for whole mount before staining with F-WGA. Hyphae of Pestalotiopsis longiseta, Pseudocercospora ocellata, Botrytis cinerea and Colletotrichum theae-sinensis fluoresced brightly in whole mount and sectioned samples of infected leaf tissue. In browned tissue, hyphae did not fluoresce frequently in whole mount sample. Autofluorescence of leaf tissue was strong in browned tissue of sections, it was removed by 10% KOH treatment before staining. Penetration hyphae of C. theae-sinensis in cell wall of trichome and hyphae in basal part of trichome did not fluoresced frequently. In whole mount samples of tea leaf infected with Exobasidium vexans and E. reticulatum, hymenia appeared on leaf surface fluoresced, but hyphae in leaf tissue did not fluoresce. In sectioned samples, hyphae fluoresced brightly when sections were treated with 10% KOH before staining.

Distinction between papillary thyroid hyperplasia and papillary thyroid carcinoma by immunohistochemical staining for cytokeratin 19, galectin-3, and HBME-1.

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Casey, Mary B; Lohse, Christine M; Lloyd, Ricardo V

2003-01-01

The histopathology of papillary thyroid hyperplasia and papillary thyroid carcinoma is similar enough to cause a diagnostic dilemma in a few cases. Both lesions may have papillary fronds with fibrovascular cores, nuclear crowding, and nuclear anisocytosis. Formalin- fixed paraffin-embedded tissues from 30 randomly selected patients with papillary thyroid hyperplasia and an equal number from patients with papillary thyroid carcinoma were analyzed for expression of cytokeratin 19 (CK19), galectin-3, and HBME-1. Cases of papillary thyroid carcinoma had moderate to strong CK19, galectin-3, and HBME-1 reactivity although both CK19 and galectin-3 showed positive staining in a significant number of nonneoplastic thyroid cases. HBME-1 was uncommon in the nonneoplastic cases. These results indicate that HBME-1 may be useful in helping to distinguish papillary thyroid carcinoma from hyperplasia in diagnostically difficult cases.

Immunohistochemical proliferation markers may overestimate the growth potential after ionizing radiation. In vivo study in the rat anterior pituitary gland

International Nuclear Information System (INIS)

Nakasu, Satoshi; Fukami, Tadateru; Matsuda, Masayuki; Nakasu, Yoko

2003-01-01

The effect of ionizing radiation on the expression of immunohistochemical proliferation markers was examined in the rat pituitary gland. Rats were irradiated in the pituitary region with a dose of 40 Gy, or were sham-irradiated as controls. Bromodeoxyuridine (BrdU) was given to the rats after one week, either one hour (Br-1 group) or 17 hours (Br-17 group) before perfusion fixation. Immunohistochemical staining for BrdU, topoisomerase II-alpha (TopoII), Ki-67 (MIB-5), p21 WAF1/CiP1 (p21), and p27 Kip1 (p27) was performed. Apoptotic cells were detected by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling method. The mean BrdU labeling index (LI) and MIB-5 LI were significantly higher in the irradiated rats than in the sham rats in the Br-1 group. TopoII LI was higher in the irradiated rats than in the sham rats, although not significantly. p27-positive cells decreased in irradiated rats, but p21-positive cells increased more than in the sham rats. The number of apoptotic cells increased significantly after radiation. BrdU LIs were lower in the irradiated rats than in the sham rats in the Br-17 group. A few small BrdU-positive fragments with apoptotic features were phagocytosed in the anterior lobe cells. These results indicate that some ''immunohistochemically proliferating cells'' subsequently undergo apoptosis in the irradiated pituitary gland. The values of proliferative indices should be cautiously interpreted after irradiation of tissue. (author)

[Parasitological, immunohistochemical and histopathological study for Leishmania chagasi detection in splenic tissues of dogs with visceral leishmaniasis].

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Tasca, Karen Ingrid; Buzetti, Wilma Aparecida Starke; Tenorio, Michely da Silva; Paulan, Silvana de Cássia; Lima, Flávia Luna; de Queiroz, Nina Mari Gual Pimenta; Machado, Rosângela Zacarias; Oliveira, Tricia Maria Ferreira de Souza; Neves, Maria Francisca; de Noronha, Antonio Carlos Faconti; de Assis, Juliana

2009-01-01

The purpose of this work was a Canine Visceral Leishmaniasis--CVL study by parasitological direct examination of Leishmania (L.) chagasi (imprinting and histological), immunohistochemical test and histopathological analysis using spleen tissues from 34 dogs euthanized by the Zoonotic Disease Control Centre from Ilha Solteira, SP, Brazil. According to the clinical signs, the dogs were divided in three groups: asymptomatics (8 dogs), oligosymptomatics (17 dogs) and symptomatics (9 dogs). After the accomplishment of all diagnostic tests, 22 dogs were considered positives (64.7%) and 12 (35.3%) were negatives to CVL. From these positive dogs, 1/22 (4.5%) was asymptomatic, 12/22(54.5%) were oligosymptomatics and 8/22 (40.1%) were symptomatics. The histopathological study in spleen tissues from positive, especially symptomatic dogs, showed a diffuse chronic inflammation with thickness of capsular and trabecular regions and there was extensive morphologic alteration of the red and white pulp by the presence of abundant macrophages full with amastigotes, the granulomatous inflammatory reaction and haemorrhagic areas. The data of this work from histopathologic examination and direct microscopic visualization of L. (L.) chagasi showed that the spleen was an useful organ to collect sample tissues for CVL diagnosis. The immunostaining detected the highest number of positive dogs and were considered an important and conclusive method to be used in addition to parasitological methods for CVL, particularly in asymptomatic or oligosymptomatic dogs.

Detection of mitotic figures in thin melanomas--immunohistochemistry does not replace the careful search for mitotic figures in hematoxylin-eosin stain.

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Ottmann, Karl; Tronnier, Michael; Mitteldorf, Christina

2015-10-01

The mitotic rate is an important prognostic criterion in patients with thin melanoma ≤ 1 mm. The aim of this study was to investigate the reproducibility of the mitotic rate in thin melanoma in hematoxylin-eosin (H&E) stain and compare it with the detection of mitotic figures by immunohistochemistry. The number of mitoses stated in the routine diagnostic report in 190 pT1 melanomas was compared with the number gained from re-evaluation of H&E sections and the number detected after staining with the mitotic marker, phosphohistone H3 (PHH3). Two different approaches were used for choosing the "hot spot" for evaluation (dermal vs epidermal/dermal). Comparing routine H&E-stained slides with re-evaluation slides, the number of mitotic figures was slightly variable. However, findings did not result in a change of the tumor stage. In 34% of the tumors with dermal mitotic figures on H&E, mitoses could not be found in the corresponding PHH3 slide anymore. In 4% of the cases, stage relevant mitoses could only be found by PHH3 immunohistochemistry. This is a single center study. Immunohistochemical staining for mitotic figures does not replace a careful evaluation of H&E-stained slides. Immunohistochemical detection of mitosis is only an additional tool; the time-saving effect is therefore negligible. Copyright © 2015 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.

Immunohistochemical and Morphofunctional Studies of Skeletal Muscle Tissues with Electric Nerve Stimulation by In Vivo Cryotechnique

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Fukasawa, Yuki; Ohno, Nobuhiko; Saitoh, Yurika; Saigusa, Takeshi; Arita, Jun; Ohno, Shinichi

2015-01-01

In this study, morphological and immunohistochemical alterations of skeletal muscle tissues during persistent contraction were examined by in vivo cryotechnique (IVCT). Contraction of gastrocnemius muscles was induced by sciatic nerve stimulation. The IVCT was performed immediately, 3 min or 10 min after the stimulation start. Prominent ripples of muscle fibers or wavy deformation of sarcolemma were detected immediately after the stimulation, but they gradually diminished to normal levels during the stimulation. The relative ratio of sarcomere and A band lengths was the highest in the control group, but it immediately decreased to the lowest level and then gradually recovered at 3 min or 10 min. Although histochemical intensity of PAS reaction was almost homogeneous in muscle tissues of the control group or immediately after the stimulation, it decreased at 3 min or 10 min. Serum albumin was immunolocalized as dot-like patterns within some muscle fibers at 3 min stimulation. These patterns became more prominent at 10 min, and the dots got larger and saccular in some sarcoplasmic regions. However, IgG1 and IgM were immunolocalized in blood vessels under nerve stimulation conditions. Therefore, IVCT was useful to capture the morphofunctional and metabolic changes of heterogeneous muscle fibers during the persistent contraction

Cartilaginous choristoma of the tongue with an immunohistochemical study

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Pereira, Gregório Wrublevski; Pereira, Volnei David; Pereira Junior, José Antonio de Castro; da Silva, Rosemeri Maurici

2012-01-01

By definition, choristomas are normal tissues found in anomalous topography. The cartilaginous features of these lesions are rare in the soft tissues of the oral cavity. The majority of cartilaginous choristomas of the tongue—the primary site of emergence of the oropharynx—are associated with adipose, fibrous or bone tissues—apart from that, only a few of these were confirmed by an immunohistochemical study. The neoplasm exclusively composed of chondromatous tissue is extremely rare in the tongue. This paper reports the clinical, surgical and pathological characteristics of a cartilaginous choristoma of the tongue diagnosed in a 64-year-old woman. PMID:23220826

In vitro cartilage tissue engineering using cancellous bone matrix gelatin as a biodegradable scaffold

International Nuclear Information System (INIS)

Yang Bo; Yin Zhanhai; Cao Junling; Shi Zhongli; Zhang Zengtie; Liu Fuqiang; Song Hongxing; Caterson, Bruce

2010-01-01

In this study, we constructed tissue-engineered cartilage using allogeneic cancellous bone matrix gelatin (BMG) as a scaffold. Allogeneic BMG was prepared by sequential defatting, demineralization and denaturation. Isolated rabbit chondrocytes were seeded onto allogeneic cancellous BMG, and cell-BMG constructs were harvested after 1, 3 and 6 weeks for evaluation by hematoxylin and eosin staining for overall morphology, toluidine blue for extracellular matrix (ECM) proteoglycans, immunohistochemical staining for collagen type II and a transmission electron microscope for examining cellular microstructure on BMG. The prepared BMG was highly porous with mechanical strength adjustable by duration of demineralization and was easily trimmed for tissue repair. Cancellous BMG showed favorable porosity for cell habitation and metabolism material exchange with larger pore sizes (100-500 μm) than in cortical BMG (5-15 μm), allowing cell penetration. Cancellous BMG also showed good biocompatibility, which supported chondrocyte proliferation and sustained their differentiated phenotype in culture for up to 6 weeks. Rich and evenly distributed cartilage ECM proteoglycans and collagen type II were observed around chondrocytes on the surface and inside the pores throughout the cancellous BMG. Considering the large supply of banked bone allografts and relatively convenient preparation, our study suggests that allogeneic cancellous BMG is a promising scaffold for cartilage tissue engineering.

In vitro cartilage tissue engineering using cancellous bone matrix gelatin as a biodegradable scaffold

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Yang Bo; Yin Zhanhai; Cao Junling; Shi Zhongli; Zhang Zengtie; Liu Fuqiang [College of Medicine, Xi' an Jiaotong University, Yanta West Road, No 76, Yanta District, Xi' an, Shaanxi Province 710061 (China); Song Hongxing [Department of Orthopedics, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Caterson, Bruce, E-mail: caojl@mail.xjtu.edu.c [Connective Tissue Biology Laboratories, Cardiff School of Biosciences, Cardiff University, Biomedical Building, Museum Avenue, Cardiff, CF10 3US (United Kingdom)

2010-08-01

In this study, we constructed tissue-engineered cartilage using allogeneic cancellous bone matrix gelatin (BMG) as a scaffold. Allogeneic BMG was prepared by sequential defatting, demineralization and denaturation. Isolated rabbit chondrocytes were seeded onto allogeneic cancellous BMG, and cell-BMG constructs were harvested after 1, 3 and 6 weeks for evaluation by hematoxylin and eosin staining for overall morphology, toluidine blue for extracellular matrix (ECM) proteoglycans, immunohistochemical staining for collagen type II and a transmission electron microscope for examining cellular microstructure on BMG. The prepared BMG was highly porous with mechanical strength adjustable by duration of demineralization and was easily trimmed for tissue repair. Cancellous BMG showed favorable porosity for cell habitation and metabolism material exchange with larger pore sizes (100-500 {mu}m) than in cortical BMG (5-15 {mu}m), allowing cell penetration. Cancellous BMG also showed good biocompatibility, which supported chondrocyte proliferation and sustained their differentiated phenotype in culture for up to 6 weeks. Rich and evenly distributed cartilage ECM proteoglycans and collagen type II were observed around chondrocytes on the surface and inside the pores throughout the cancellous BMG. Considering the large supply of banked bone allografts and relatively convenient preparation, our study suggests that allogeneic cancellous BMG is a promising scaffold for cartilage tissue engineering.

Immunohistochemical expression of p53 proteins in Wilms' tumour: a possible association with the histological prognostic parameter of anaplasia.

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Cheah, P L; Looi, L M; Chan, L L

1996-01-01

Wilms' tumour (nephroblastoma) has been associated with chromosomal abnormalities at the 11p13, 11p15 and 16q regions. A study into the possibility of mutations occurring within p53, the ubiquitous adult tumour suppressor gene, in Wilms' tumour was carried out. Thirty-eight cases were studied. Of these 36 were categorised into the favourable histology group and two into the unfavourable histology group based on the National Wilms' Tumour Study criteria. Archival formalin-fixed, paraffin-embedded tissue sections from each case were stained with a polyclonal (AB565:Chemicon) and a monoclonal (DO7:Dako) antibody raised against p53 protein using a peroxidase-labelled streptavidin biotin kit (Dako). 'Cure' (disease-free survival of 60 months or longer) was documented in 39% of cases with favourable histology tumours. Eleven percent in this group succumbed to the disease. Both cases with unfavourable histology died. Four out of 36 (11%) tumours with favourable histology demonstrated weak to moderate staining with both AB565 and DO7 in more than 75% of tumour cells. In contrast, p53 protein expression in unfavourable histology tumours was significantly increased compared with the favourable histology group (P = 0.021) with both cases demonstrating immunopositivity in > 75% of tumour cells when stained with AB565 and DO7. The intensity of staining ranged from moderate to strong in both cases. It appears from this preliminary study that the immunohistochemical expression of p53 protein in Wilms' tumour, presumably a result of mutation in the p53 tumour suppressor gene, correlates with histological classification, histological categorisation being one of the useful features in the prognostic assessment of Wilms' tumours.

Expression analyses of human cleft palate tissue suggest a role for osteopontin and immune related factors in palatal development

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Jakobsen, L.P.; Borup, R.; Vestergaard, J.

2009-01-01

. Moreover, selected differentially expressed genes were analyzed by quantitative RT-PCR, and by immunohistochemical staining of craniofacial tissue from human embryos. Osteopontin (SPP1) and other immune related genes were significantly higher expressed in palate tissue from patients with CLP compared to CP...... and palate (CLP). In order to understand the biological basis in these cleft lip and palate subgroups better we studied the expression profiles in human tissue from patients with CL/P. In each of the CL/P subgroups, samples were obtained from three patients and gene expression analysis was performed...... and immunostaining in palatal shelves against SPP1, chemokine receptor 4 (CXCR4) and serglycin (PRG1) in human embryonic craniofacial tissue were positive, supporting a role for these genes in palatal development. However, gene expression profiles are subject to variations during growth and therefore we recommend...

Propidium iodide (PI) stains Nissl bodies and may serve as a quick marker for total neuronal cell count.

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Niu, Junfei; Li, Chunman; Wu, Haihui; Feng, Xianling; Su, Qingning; Li, Shihe; Zhang, Lihong; Yew, David Tai Wai; Cho, Eric Yu Pang; Sha, Ou

2015-03-01

Propidium iodide (PI) reacts with both DNA and RNA and is a commonly used fluorescent reagent for nucleic acid staining. The aim of the study was to compare the cellular staining patterns of PI with that of Nissl staining in rat nervous tissues and to report a modified staining method that selectively labels Nissl bodies in neurons. Cryosections and paraffin sections of different tissues of normal Sprague-Dawley rats, including trigeminal ganglia, dorsal root ganglia, spinal cord, liver, and small intestine, were stained by either PI or the hematoxylin and eosin method. Some sections were treated with RNase or DNase before the above staining, and some were double stained with PI and a Nissl stain. The sections were observed by light, fluorescence or confocal microscopy. Results showed strong PI signals detected as patterns of granules in the neuronal cytoplasm of all nervous tissues, whereas the staining of neuronal nuclei was weaker. In contrast, nuclei of neuroglial cells were strongly stained by PI, while the cytoplasm was not obviously stained. Pretreatment of the neural tissue with RNase abolished the PI signals. Furthermore, the PI positive granules in neuronal cytoplasm co-localized with Nissl bodies stained by the fluorescent Nissl stain. When the tissue was pretreated with DNase, PI only stained the cytoplasmic granules of neurons, but not that of glial cells. Our results show that PI stains Nissl bodies and may serve as an economical and convenient neuron marker for neuronal cell counting when specific neural markers such as antibodies are not readily available. Copyright © 2015. Published by Elsevier GmbH.

Immunohistochemical evaluation of myofibroblasts in odontogenic cysts and tumors: A comparative study.

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Syamala, Deepa; Suresh, Rakesh; Janardhanan, Mahija; Savithri, Vindhya; Anand, Prem P; Jose, Amrutha

2016-01-01

Myofibroblasts are fibroblasts with smooth muscle-like features characterized by the presence of a contractile apparatus and found in the connective tissue stroma of normal tissues such as blood vessels and lymph nodes. They are now thought to play a role in the synthesis and reorganization of extracellular matrix, which could contribute to the aggressive biologic behavior of the lesions. To compare the mean number of stromal myofibroblasts in dentigerous cysts (DCs), keratocystic odontogenic tumor (KCOT) and ameloblastoma; and to derive a correlation between the stromal myofibroblasts and the known biologic behavior of the lesions. A cross-sectional immunohistochemical analysis of cases of DC, KCOT and ameloblastoma. Twenty paraffin-embedded tissue blocks each of DC, KCOT and multicystic ameloblastoma were selected for the study and diagnosis confirmed through hematoxylin and eosin staining. Tissue sections were analyzed for the number of myofibroblasts using alpha smooth muscle actin (α-SMA) immunostaining. Differences in the mean number of α-SMA positive cells in each group were analyzed using one-way ANOVA test. Intergroup comparisons of mean values of α-SMA positive cells were performed using Mann-Whitney U-test. Ameloblastoma showed the highest number of myofibroblasts, whereas DC showed the lowest. Among the groups, there were significant differences between the myofibroblast counts among DC and KCOT and between DC and ameloblastoma, whereas the difference in counts was not statistically significant between KCOT and ameloblastoma. A positive correlation was observed between the myofibroblast count and the known biologic behavior of the lesions. Myofibroblasts may act in close association with the epithelial cells to bring about changes in stromal microenvironment, favorable to the growth and progression of the lesion. They may be of great value in predicting the biologic behavior and growth potential of such lesions.

Immunohistochemical Ki-67/KL1 double stains increase accuracy of Ki-67 indices in breast cancer and simplify automated image analysis

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Nielsen, Patricia S; Bentzer, Nina K; Jensen, Vibeke

2014-01-01

observers and automated image analysis. RESULTS: Indices were predominantly higher for single stains than double stains (P≤0.002), yet the difference between observers was statistically significant (PPearson correlation coefficient for manual and automated indices ranged from 0.......69 to 0.85 (Pcorrelating automated indices with tumor characteristics, for example, tumor size (P... stains, Ki-67 should be quantified on double stains to reach a higher accuracy. Automated indices correlated well with manual estimates and tumor characteristics, and they are thus possibly valuable tools in future exploration of Ki-67 in breast cancer....

Histopathological evaluation of ocular microsporidiosis by different stains

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Sharma Savitri

2006-06-01

Full Text Available Abstract Background There is limited data on comparing stains in the detection of microsporidia in corneal biopsies. Hence we wanted to evaluate various stains for their ability to detect microsporidia in corneal tissue sections. Methods Four cases diagnosed with microsporidiosis on Hematoxylin and Eosin and Periodic Acid Schiff's stained sections of the corneal button between January 2002 and December 2004, were included. Further sections were prospectively stained with calcofluor white, Gram, Giemsa, Masson's trichrome, acridine orange, Gomori's methenamine silver, Gram's chromotrope and modified acid fast stain. The stained sections were analyzed for the spore characteristics in terms of size, shape, color contrast, cell wall morphology, waist band in cytoplasm and ease of detection. Results All sections showed microsporidial spores as 3 – 5 μm, oval bodies. 1% acid fast, Gram's chromotrope and GMS stains provided a reliable diagnosis of microsporidia as diagnostic waist band could be identified and good contrast helped distinguish the spores from inflammatory debris. Conclusion Considering the ease of performance, cost effectiveness and rapidity of the technique, 1% acid fast stain and Gram's chromotrope stain are ideal for the detection of microsporidia.

Twin Rectal Tonsils Mimicking Carcinoid or Mucosa-Associated Lymphoid Tissue Lymphoma.

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Takehara, Masanori; Muguruma, Naoki; Kitamura, Shinji; Kimura, Tetsuo; Okamoto, Koichi; Miyamoto, Hiroshi; Bando, Yoshimi; Takayama, Tetsuji

2017-09-01

The rectal tonsil is a rare polypoid lesion exclusively found in the rectum and is considered a reactive proliferation of the lymphoid tissue. Although this lesion is benign, we recommend that it should be differentiated from carcinoid or polypoid type of mucosa-associated lymphoid tissue lymphomas, based on gross findings. In this case report, we describe a case of rectal lesions with a unique appearance in a 41-year-old man. Colonoscopy revealed two 5-mm-sized nodules located opposite from each other on the left and right sides of the lower rectum. Endoscopic mucosal resection was conducted. Histopathologically, both lesions were mainly located in the submucosa and consisted of prominent lymphoid follicles with germinal centers of various sizes. No immunoreactivity of Bcl-2 was seen in the germinal centers. Immunohistochemical staining for kappa and lambda light chains revealed a polyclonal pattern. Therefore, these lesions were diagnosed as rectal tonsils.

Histological and Immunohistochemical Revision of Hepatocellular Adenomas: A Learning Experience

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S. Fonseca

2013-01-01

Full Text Available Light has been shed on the genotype/phenotype correlation in hepatocellular adenoma (HCA recognizing HNF1α-inactivated HCA (H-HCA, inflammatory HCA (IHCA, and β-catenin-activated HCA (b-HCA. We reviewed retrospectively our surgical HCA series to learn how to recognize the different subtypes histopathologically and how to interpret adequately their immunohistochemical staining. From January 1992 to January 2012, 37 patients underwent surgical resection for HCA in our institution. Nine had H-HCA (25% characterized by steatosis and loss of L-FABP expression; 20 had IHCA (55.5% showing CRP and/or SAA expression, sinusoidal dilatation, and variable inflammation; and 1 patient had both H-HCA and IHCA. In 5 patients (14%, b-HCA with GS and β-catenin nuclear positivity was diagnosed, two already with hepatocellular carcinoma. Two cases (5.5% remained unclassified. One of the b-HCA showed also the H-HCA histological and immunohistochemical characteristics suggesting a subgroup of β-catenin-activated/HNF1α-inactivated HCA, another b-HCA exhibited the IHCA histological and immunohistochemical characteristics suggesting a subgroup of β-catenin-activated/inflammatory HCA. Interestingly, three patients had underlying vascular abnormalities. Using the recently published criteria enabled us to classify histopathologically our retrospective HCA surgical series with accurate recognition of b-HCA for which we confirm the higher risk of malignant transformation. We also underlined the association between HCA and vascular abnormalities.

 Immunohistochemical Expression of ki-67 and p53 in Colorectal Adenomas: A Clinicopathological Study

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Hussam Hasson Ali

2011-07-01

Full Text Available  Objectives: To evaluate the significance of P53 and Ki-67 expression as immunohistochemical markers in early detection of premalignant changes in different types of colorectal adenomas. Also, to correlate immunohistochemical expression of the two markers with different clinicopathological parameters including; age, and sex of the patient, type, site, size and grade of dysplasia of colorectal adenomas.Methods: Forty-seven polypectomy specimens of colorectal adenomas were retrieved from the archival materials of the Gastrointestinal and Hepatic Diseases Teaching Hospital in Baghdad from 2009 - 2010. Four µm section specimens were stained by immunohistochemical technique with Ki-67 and P53 tumor markers. P-values <0.05 were considered statistically significant.Results: Immunohistochemical expressions of Ki-67 and P53 had a significant correlation with the size and grade of dysplasia in colorectal adenomas. However, there was no significant correlation among the immunohistochemical expression of Ki-67 and P53 with the age and gender of the patient, and the type and site of colorectal adenomas. There was no significant correlation between Ki-67 and P53 expressions in colorectal adenomas. Villous adenomas of colorectum showed a significant correlation with the grade of dysplasia, while there was no significant correlation between size and site of colorectal adenoma with the grade of dysplasia.Conclusion: High grade dysplasia with significant positive immunohistochemical markers of Ki-67 and P53 could be valuable parameters for selecting from the total colorectal adenoma population, those most deserving of close surveillance in follow-up cancer prevention programs. It is closely linked with increasing age particularly in patients with a large size adenoma of villous component in their histology.

Evaluation of a panel of antibodies for the immunohistochemical identification of immune cells in paraffin-embedded lymphoid tissues of new- and old-world camelids.

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Uhde, Ann-Kathrin; Lehmbecker, Annika; Baumgärtner, Wolfgang; Spitzbarth, Ingo

2017-02-01

Different species of camelids play an important role in the epidemiology of various emerging infectious diseases such as Middle East respiratory syndrome. For precise investigations of the immunopathogenesis in these host species, appropriate immunohistochemical markers are highly needed in order to phenotype distinct immune cells populations in camelids. So far, specific immunohistochemical markers for camelid immune cells are rarely commercially available, and cross-reactivity studies are restricted to the use of frozen dromedary tissues. To bridge this gap, 14 commercially available primary antibodies were tested for their suitability to demonstrate immune cell populations on formalin fixed paraffin-embedded (FFPE) tissue sections of dromedaries, Bactrian camels, llamas, and alpacas in the present study. Out of these, 9 antibodies directed against CD3, CD20, CD79α, HLA-DR, Iba-1, myeloid/histiocyte antigen, CD204, CD208, and CD68 antigen exhibited distinct immunoreaction patterns to certain camelid immune cell subsets. The distribution of these antigens was comparatively evaluated in different anatomical compartments of thymus, spleen, mesenteric, and tracheobronchial lymph nodes. The presented results will provide a basis for further investigations in camelids, especially with respect to the role of the immune response in certain infectious diseases, which harbor a considerable risk to spill over to other species. Copyright © 2017 Elsevier B.V. All rights reserved.

Cellular and Subcellular Immunohistochemical Localization and Quantification of Cadmium Ions in Wheat (Triticum aestivum.

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Wei Gao

Full Text Available The distribution of metallic ions in plant tissues is associated with their toxicity and is important for understanding mechanisms of toxicity tolerance. A quantitative histochemical method can help advance knowledge of cellular and subcellular localization and distribution of heavy metals in plant tissues. An immunohistochemical (IHC imaging method for cadmium ions (Cd2+ was developed for the first time for the wheat Triticum aestivum grown in Cd2+-fortified soils. Also, 1-(4-Isothiocyanobenzyl-ethylenediamine-N,N,N,N-tetraacetic acid (ITCB-EDTA was used to chelate the mobile Cd2+. The ITCB-EDTA/Cd2+ complex was fixed with proteins in situ via the isothiocyano group. A new Cd2+-EDTA specific monoclonal antibody, 4F3B6D9A1, was used to locate the Cd2+-EDTA protein complex. After staining, the fluorescence intensities of sections of Cd2+-positive roots were compared with those of Cd2+-negative roots under a laser confocal scanning microscope, and the location of colloidal gold particles was determined with a transmission electron microscope. The results enable quantification of the Cd2+ content in plant tissues and illustrate Cd2+ translocation and cellular and subcellular responses of T. aestivum to Cd2+ stress. Compared to the conventional metal-S coprecipitation histochemical method, this new IHC method is quantitative, more specific and has less background interference. The subcellular location of Cd2+ was also confirmed with energy-dispersive X-ray microanalysis. The IHC method is suitable for locating and quantifying Cd2+ in plant tissues and can be extended to other heavy metallic ions.

Cellular and Subcellular Immunohistochemical Localization and Quantification of Cadmium Ions in Wheat (Triticum aestivum).

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Gao, Wei; Nan, Tiegui; Tan, Guiyu; Zhao, Hongwei; Tan, Weiming; Meng, Fanyun; Li, Zhaohu; Li, Qing X; Wang, Baomin

2015-01-01

The distribution of metallic ions in plant tissues is associated with their toxicity and is important for understanding mechanisms of toxicity tolerance. A quantitative histochemical method can help advance knowledge of cellular and subcellular localization and distribution of heavy metals in plant tissues. An immunohistochemical (IHC) imaging method for cadmium ions (Cd2+) was developed for the first time for the wheat Triticum aestivum grown in Cd2+-fortified soils. Also, 1-(4-Isothiocyanobenzyl)-ethylenediamine-N,N,N,N-tetraacetic acid (ITCB-EDTA) was used to chelate the mobile Cd2+. The ITCB-EDTA/Cd2+ complex was fixed with proteins in situ via the isothiocyano group. A new Cd2+-EDTA specific monoclonal antibody, 4F3B6D9A1, was used to locate the Cd2+-EDTA protein complex. After staining, the fluorescence intensities of sections of Cd2+-positive roots were compared with those of Cd2+-negative roots under a laser confocal scanning microscope, and the location of colloidal gold particles was determined with a transmission electron microscope. The results enable quantification of the Cd2+ content in plant tissues and illustrate Cd2+ translocation and cellular and subcellular responses of T. aestivum to Cd2+ stress. Compared to the conventional metal-S coprecipitation histochemical method, this new IHC method is quantitative, more specific and has less background interference. The subcellular location of Cd2+ was also confirmed with energy-dispersive X-ray microanalysis. The IHC method is suitable for locating and quantifying Cd2+ in plant tissues and can be extended to other heavy metallic ions.

Immunohistochemical Expression of p53 in Pleomorphic Adenoma and Carcinoma Ex Pleomorphic Adenoma

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Tarakji, B.; Kujan, O.; Nassani, M. Z.

2010-01-01

Context. Immunohistochemical stains for p53 are used as a diagnostic marker associated with malignancy in several histologic types of salivary gland tumors. This marker may be useful in differentiating pleomorphic adenoma (PA) from carcinoma ex pleomorphic adenoma (CPA), as these tumors are often difficult to distinguish on the basis of morphology alone. Objective. to evaluate whatever inactivation of tumor suppressor gene (p53) increases with the tumor progression from normal salivary tissue to PA and eventually CPA. Design. Paraffin blocks of 29 cases of PA, which were surrounded by normal parotid gland, and 27 cases of carcinoma ex pleomorphic adenoma were retrieved and validated. In all cases of carcinoma ex pleomorphic adenoma, a PA “ghost” was identified, and the malignant element was either undifferentiated carcinoma or adenocarcinoma. Results. The results showed negative nuclear expression of P53 in normal parotid gland. Nuclear P53 was expressed strongly in 6/29 (20.7%) pleomorphic salivary adenoma and 10/27 (37%) carcinoma ex pleomorphic adenoma. Conclusion. Our data suggest that inactivation of p53 may play an important role in the evolution of pleomorphic salivary adenoma and carcinoma ex pleomorphic adenoma.

Tissue-engineered cartilaginous constructs for the treatment of caprine cartilage defects, including distribution of laminin and type IV collagen.

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Jeng, Lily; Hsu, Hu-Ping; Spector, Myron

2013-10-01

The purpose of this study was the immunohistochemical evaluation of (1) cartilage tissue-engineered constructs; and (2) the tissue filling cartilage defects in a goat model into which the constructs were implanted, particularly for the presence of the basement membrane molecules, laminin and type IV collagen. Basement membrane molecules are localized to the pericellular matrix in normal adult articular cartilage, but have not been examined in tissue-engineered constructs cultured in vitro or in tissue filling cartilage defects into which the constructs were implanted. Cartilaginous constructs were engineered in vitro using caprine chondrocyte-seeded type II collagen scaffolds. Autologous constructs were implanted into 4-mm-diameter defects created to the tidemark in the trochlear groove in the knee joints of skeletally mature goats. Eight weeks after implantation, the animals were sacrificed. Constructs underwent immunohistochemical and histomorphometric evaluation. Widespread staining for the two basement membrane molecules was observed throughout the extracellular matrix of in vitro and in vivo samples in a distribution unlike that previously reported for cartilage. At sacrifice, 70% of the defect site was filled with reparative tissue, which consisted largely of fibrous tissue and some fibrocartilage, with over 70% of the reparative tissue bonded to the adjacent host tissue. A novel finding of this study was the observation of laminin and type IV collagen in in vitro engineered cartilaginous constructs and in vivo cartilage repair samples from defects into which the constructs were implanted, as well as in normal caprine articular cartilage. Future work is needed to elucidate the role of basement membrane molecules during cartilage repair and regeneration.

A robust, efficient and flexible method for staining myelinated axons in blocks of brain tissue.

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Wahlsten, Douglas; Colbourne, Frederick; Pleus, Richard

2003-03-15

Previous studies have demonstrated the utility of the gold chloride method for en bloc staining of a bisected brain in mice and rats. The present study explores several variations in the method, assesses its reliability, and extends the limits of its application. We conclude that the method is very efficient, highly robust, sufficiently accurate for most purposes, and adaptable to many morphometric measures. We obtained acceptable staining of commissures in every brain, despite a wide variety of fixation methods. One-half could be stained 24 h after the brain was extracted and the other half could be stained months later. When staining failed because of an exhausted solution, the brain could be stained successfully in fresh solution. Relatively small changes were found in the sizes of commissures several weeks after initial fixation or staining. A half brain stained to reveal the mid-sagittal section could then be sectioned coronally and stained again in either gold chloride for myelin or cresyl violet for Nissl substance. Uncertainty, arising from pixelation of digitized images was far less than errors arising from human judgments about the histological limits of major commissures. Useful data for morphometric analysis were obtained by scanning the surface of a gold chloride stained block of brain with an inexpensive flatbed scanner.

Immunohistochemical investigations of xenotransplanted human adenocarcinomas on nude mice: Correlation to radioimaging

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Matejkova, E.

1987-01-01

Immunohistochemical investigations of xenotransplanted human adenocarcinomas on nude mice; correlation to radioimaging Human carcinomas were subcutanously grafted to nude mice (Balb/c-nu/nu) and were investigated in four passages by immunohistochemical methods and by the fluorochrome bisbenzimid. In this way there could be observed a successful differentiation between the nourishing murine stroma and the human tumor parenchym. Especially the use of a monoclonal antibody (rat/mouse fusion) directed against human tissue turned out to be a suitable method. Four adenocarcinomas were tested: Colon-, mamma-, stomach- and testicle carcinoma. During the first four passages atypical parts of murine connective tissue and some changes in the human parenchyma could be seen. These results demonstrate that also in nude mice variations of the transplanted tumor material could happen. They could be detected in time with a routine immunohistochemical test. The consequences of tumor morphological variations for the development of therapeutic and diagnostic tools were studied with the help of radioimaging by external scintigraphy. Furthermore the biodistribution, tumoruptake and the whole body counting were studied by means of radionuclid marked monoclonal antibodies. The morphological variations of the passages of mammary, testicle and colon carcinomas were not big enough to influence the results in a certain way. Therefore especially the relation between the activity uptake in the tissue, the size of the tumor and the whole body uptake was studied in view of immunoscintigraphy. (orig./MG) [de

A single simple procedure for dewaxing, hydration and heat-induced epitope retrieval (HIER for immunohistochemistry in formalin fixed paraffin-embedded tissue

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I.M.S. Paulsen

2015-11-01

Full Text Available Heat-induced epitope retrieval (HIER is widely used for immunohistochemistry on formalin fixed paraffin-embedded tissue and includes temperatures well above the melting point of paraffin. We therefore tested whether traditional xylene-based removal of paraffin is required on sections from paraffin-embedded tissue, when HIER is performed by vigorous boiling in 10 mM Tris/0.5 mM EGTA-buffer (pH=9. Immunohistochemical results using HIER with or without prior dewaxing in xylene were evaluated using 7 primary antibodies targeting proteins located in the cytosol, intracellular vesicles and plasma membrane. No effect of omitting prior dewaxing was observed on staining pattern. Semiquantitative analysis did not show HIER to influence the intensity of labelling consistently. Consequently, quantification of immune labelling intensity using fluorescent secondary antibodies was performed at 5 dilutions of primary antibody with and without prior dewaxing in xylene. No effect of omitting prior dewaxing on signal intensity was detectable indicating similar immunoreactivity in dewaxed and non-dewaxed sections. The intensity of staining the nucleus with the DNA-stain ToPro3 was similarly unaffected by omission of dewaxing in xylene. In conclusion, the HIER procedure described and tested can be used as a single procedure enabling dewaxing, hydration and epitope retrieval for immunohistochemistry in formalin fixed paraffin-embedded tissue.

Diagnostic value of HMB-45 and anti-Melan A staining of sentinel lymph nodes with isolated positive cells.

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Mahmood, Muhammad N; Lee, Min W; Linden, Michael D; Nathanson, S D; Hornyak, Thomas J; Zarbo, Richard J

2002-12-01

Numerous immunohistochemical stains have been employed to detect metastatic melanoma in sentinel lymph node (SLN) biopsies. HMB-45 is considered by some as a specific tool to detect early metastatic melanoma (1). Occasionally, one or two isolated HMB-45-positive cells may cause complications in diagnostic interpretation. The goal of this study was to evaluate the reliability of HMB-45 staining of SLNs with sparse isolated positive cells and to compare its staining with anti-Melan A antibody. HMB-45 and anti-Melan A antibody immunostaining was performed on (Group A) 15 histologically negative SLNs excised from patients with malignant melanoma (MM) and on (Group B) 15 histologically negative SLNs excised from patients with breast carcinoma (BC). None of the patients had clinical evidence of systemic metastasis at the time of SLN biopsy. Five cutaneous biopsies with changes of postinflammatory hyperpigmentation (PIHP) were also stained with both antibodies. HMB-45 staining was repeated in all Group B SLNs after blocking endogenous biotins. Electron-microscopic studies were performed on all cases of PIHP. Isolated HMB-45-stained cells were present in 6 of 15 SLNs removed for MM; 8 of 15 for BC; and 3 of 5 cutaneous biopsies of PIHP. HMB-45 reactivity persisted after blocking endogenous biotins in 6 of 8 positive SLNs from Group B. Anti-Melan A antibody was negative in all SLNs of group A and B and in dermal melanophages of all five cases of PIHP. HMB-45 positivity was demonstrated in histologically negative SLNs and cutaneous biopsies, especially in the milieu of aggregated melanophages. Phagocytosis of premelanosomes by macrophages in the draining lymph nodes may account for isolated cell positivity and can hinder correct diagnostic interpretation. HMB-45 may not be a reliable marker for the detection of micro-metastasis of MM and requires correlation with other immunohistochemical markers, such as anti-Melan A antibody, to enhance specificity.

The use of special stains in liver biopsy interpretation: Implications ...

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Materials and Methods: The formalin fixed paraffin embedded blocks of liver biopsies reported in two histopathology laboratories between 2008 and 2013 were retrieved. These were stained with H and E and the following standard special stains for liver tissue histology – Perl's Prussian blue, reticulin, Sirius red, Shikata ...

Immunohistochemical expression of CD44s in renal cell carcinoma lacks independent prognostic significance

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Walter Henriques da Costa

2012-08-01

Full Text Available PURPOSE: To analyze the immunohistochemical expression of the standard isoform of CD44 (CD44s adhesion molecule in clear cell renal cell carcinoma (CCRCC and its impact on clinical outcomes. MATERIALS AND METHODS: Ninety-nine consecutive patients treated surgically for RCC between 1992 and 2009 were selected. A single pathologist reviewed all cases to effect a uniform reclassification and determine the most representative tumor areas for construction of a tissue microarray. The same pathologist, who was blinded to the outcome of the cases, semi-quantitatively scored the staining intensity of CD44s in all specimens. The counting was done using the H-Score algorithm. RESULTS: Of the 99 immunostained RCC specimens, 57(57.7% showed low expression, and 42(42.4% showed high expression levels of CD44s. The expression of CD44s was directly associated with tumor size (p = 0.03, clinical stage (p = 0.02 and Fuhrman grade (p = 0.02. Disease specific survival (DSS rates for patients whose specimens expressed low and high levels of CD44s was 88.1% and 67.5%, respectively (p = 0.009. Progression free survival (PFS rates in patients with low and high expression of CD44s were 78.8% and 61.7%, respectively (p = 0.05. Classical features such as the presence of metastasis and clinical stage remained isolated predictors of survival. CONCLUSIONS: Immunohistochemical expression of CD44s was associated with important clinical variables such as stage and Fuhrman grade. However, it was not an independent predictor of survival. Therefore, we believe it has a limited role as a prognostic marker in patients with CCRCC.

Detection of EGFR and COX-2 Expression by Immunohistochemical Method on a Tissue Microarray Section in Lung Cancer and Biological Significance

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Xinyun WANG

2010-02-01

Full Text Available Background and objective Epidermal growth factor receptor (EGFR and cyclooxygenase-2 (COX-2, which can regulate growth, invasion and metastasis of tumor through relevant signaling pathway, have been detected in a variety of solid tumors. The aim of this study is to investigate the biological significance of EGFR and COX-2 expression in lung cancer and the relationship between them. Methods The expression of EGFR and COX-2 was detected in 89 primary lung cancer tissues, 12 premaliganant lesions, 12 lymph node metastases, and 10 normal lung tissues as the control by immunohistochemical method on a tissue microarray section. Results EGFR protein was detectable in 59.6%, 41.7%, and 66.7% of primary lung cancer tissues, premalignant lesions and lymph node metastases, respectively; COX-2 protein was detectable in 52.8%, 41.7%, and 66.7% of primary lung cancer tissues, premalignant lesions and lymph node metastases, respectively, which were significantly higher than those of the control (P 0.05. COX-2 expression was related to gross type (P < 0.05. A highly positive correlation was observed between EGFR and COX-2 expression (P < 0.01. Conclusion Overexpression of EGFR and COX-2 may play an important role in the tumorgenesis, progression and malignancy of lung cancer. Detection of EGFR and COX-2 expression might be helpful to diagnosis and prognosis of lung cancer.

Histopathological and immunohistochemical analysis of small intestinal biopsies in adults suspected of celiac disease

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Iftikhar, R.; Jamal, S.; Zafar, A.; Saadia, A.

2016-01-01

Objective: To analyse histomorphological and immunohistochemical analysis of small intestinal biopsies in adults suspected of celiac disease. Study Design: Descriptive study. Place and Duration of Study: Department of Histopathology, Army Medical College, Rawalpindi, from November 2014 to December 2015. Methodology: Fifty cases of small intestinal mucosal biopsies (duodenal and jejunal) were analysed in adult patients aged above 14 years suspected of celiac disease. Their histomorphological data was recorded using Modified Marsh Criteria. Type of intraepithelial lymphocytes was assessed using immunohistochemistry. Intraepithelial lymphocytes were counted both by H and E stain and immunostain CD3 and CD20. Results: Thirty-four percent patients were aged between 21 - 30 years and 22% patients aged between 41 - 50 years. There were 84% (n=42) males. Thirteen (26%) cases showed focal villous atrophy, 32 (64%) cases showed partial villous atrophy and 5 (10%) cases showed complete villous atrophy. Anti-tissue transglutaminase antibody was positive in 21 (42%) cases. CD3 immunomarker was positive for intraepithelial lymphocytes in all 50 cases while CD20 immunomarker showed focal positivity in areas with lymphoid follicle formation. The count of intraepithelial lymphocytes was found to be almost equal (with a difference of 3 - 4 lymphocytes) on both H and E stain and immunostain CD3 and CD20. Conclusion: Males aged 21 - 30 years were the most commonly affected group. The most frequent change in histology was partial villous atrophy along with lymphocytic enteritis. All the intraepithelial lymphocytes were present in crescendo-pattern of distribution. (author)

CARCINOMA PROSTATE HISTOPATHOLOGY IN NEEDLE BIOPSIES INCLUDING REVISED GLEASON’S GRADING AND ROLE OF IMMUNOHISTOCHEMICAL MARKERS

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Rema Priyadarsini

2017-05-01

Full Text Available BACKGROUND Adenocarcinoma of prostate is the most common form of cancer in men accounting for 29% of cancers in developed nations and the incidence of prostatic cancer is 6.4% in males of Trivandrum District. MATERIALS AND METHODS All prostatic biopsies taken per rectally and stained by haematoxylin and eosin. In suspected cases of malignancy immunohistochemical markers, the AMACR P504S and high molecular weight cytokeratin 34E12 were done. RESULTS The total number of cases studied were 142. The final diagnosis with histomorphological features show that maximum cases were prostatic carcinoma constituting 45.5% of the samples received. CONCLUSION All prostatic carcinomas were graded by revised Gleason’s grade (ISUP 2005 and the use of immunohistochemical markers in arriving at a definite diagnosis in carcinoma prostate was confirmed.

Expression of embryonic stem cell markers in keloid-associated lymphoid tissue.

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Grant, Chelsea; Chudakova, Daria A; Itinteang, Tinte; Chibnall, Alice M; Brasch, Helen D; Davis, Paul F; Tan, Swee T

2016-07-01

To identify, characterise and localise the population of primitive cells in keloid scars (KS). 5-µm-thick formalin-fixed paraffin-embedded sections of KS samples from 10 patients underwent immunohistochemical (IHC) staining for the embryonic stem cell (ESC) markers OCT4, SOX2, pSTAT3 and NANOG, and keloid-associated lymphoid tissue (KALT) markers CD4 and CD20. NanoString gene expression analysis and in situ hybridisation (ISH) were used to determine the abundance and localisation of the mRNA for these ESC markers. IHC staining revealed the expression of the ESC markers OCT4, SOX2, pSTAT3 and NANOG by a population of cells within KS tissue. These are localised to the endothelium of the microvessels within the KALTs. NanoString gene expression analysis confirmed the abundance of the transcriptional expression of the same ESC markers. ISH localised the expression of the ESC transcripts to the primitive endothelium in KS tissue. This report demonstrates the expression of ESC markers OCT4, SOX2, pSTAT3 and NANOG by the endothelium of the microvessels within the KALTs. These findings show a unique niche of primitive cells within KS, expressing ESC markers, revealing a potential therapeutic target in the treatment of KS. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

Evidence of specialized tissue in human interatrial septum: histological, immunohistochemical and ultrastructural findings.

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Mitrofanova, Lubov B; Gorshkov, Andrey N; Lebedev, Dmitry S; Mikhaylov, Evgeny N

2014-01-01

There is a paucity of information on structural organization of muscular bundles in the interatrial septum (IAS). The aim was to investigate histologic and ultrastructural organization of muscular bundles in human IAS, including fossa ovalis (FO) and flap valve. Macroscopic and light microscopy evaluations of IAS were performed from postmortem studies of 40 patients. Twenty three IAS specimens underwent serial transverse sectioning, and 17--longitudinal sectioning. The transverse sections from 10 patients were immunolabeled for HCN4, Caveolin3 and Connexin43. IAS specimens from 6 other patients underwent electron microscopy. In all IAS specimens sections the FO, its rims and the flap valve had muscle fibers consisting of working cardiac myocytes. Besides the typical cardiomyocytes there were unusual cells: tortuous and horseshoe-shaped intertangled myocytes, small and large rounded myocytes with pale cytoplasm. The cells were aggregated in a definite structure in 38 (95%) cases, which was surrounded by fibro-fatty tissue. The height of the structure on transverse sections positively correlated with age (P = 0.03) and AF history (P = 0.045). Immunohistochemistry showed positive staining of the cells for HCN4 and Caveolin3. Electron microscopy identified cells with characteristics similar to electrical conduction cells. Specialized conduction cells in human IAS have been identified, specifically in the FO and its flap valve. The cells are aggregated in a structure, which is surrounded by fibrous and fatty tissue. Further investigations are warranted to explore electrophysiological characteristics of this structure.

Evidence of specialized tissue in human interatrial septum: histological, immunohistochemical and ultrastructural findings.

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Lubov B Mitrofanova

Full Text Available There is a paucity of information on structural organization of muscular bundles in the interatrial septum (IAS. The aim was to investigate histologic and ultrastructural organization of muscular bundles in human IAS, including fossa ovalis (FO and flap valve.Macroscopic and light microscopy evaluations of IAS were performed from postmortem studies of 40 patients. Twenty three IAS specimens underwent serial transverse sectioning, and 17--longitudinal sectioning. The transverse sections from 10 patients were immunolabeled for HCN4, Caveolin3 and Connexin43. IAS specimens from 6 other patients underwent electron microscopy.In all IAS specimens sections the FO, its rims and the flap valve had muscle fibers consisting of working cardiac myocytes. Besides the typical cardiomyocytes there were unusual cells: tortuous and horseshoe-shaped intertangled myocytes, small and large rounded myocytes with pale cytoplasm. The cells were aggregated in a definite structure in 38 (95% cases, which was surrounded by fibro-fatty tissue. The height of the structure on transverse sections positively correlated with age (P = 0.03 and AF history (P = 0.045. Immunohistochemistry showed positive staining of the cells for HCN4 and Caveolin3. Electron microscopy identified cells with characteristics similar to electrical conduction cells.Specialized conduction cells in human IAS have been identified, specifically in the FO and its flap valve. The cells are aggregated in a structure, which is surrounded by fibrous and fatty tissue. Further investigations are warranted to explore electrophysiological characteristics of this structure.

Immunohistochemical Expression of Podoplanin in Clinical Variants of Oral Leukoplakia and Its Correlation With Epithelial Dysplasia.

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D'souza, Brendan; Nayak, Ramakant; Kotrashetti, Vijayalakshmi S

2018-02-01

Oral leukoplakia is the most common premalignant lesion of the oral cavity and is associated with development of oral squamous cell carcinoma. Certain changes at cellular and molecular level are important indicators for transformation into carcinoma. Podoplanin, a transmembrane glycoprotein is involved in the cytoskeletal remodeling and increased motility of the cell which helps in determining the malignant potential of oral leukoplakic lesions. The objective of the study was to determine immunohistochemically the expression of podoplanin in homogenous (HOL) and nonhomogenous oral leukoplakia (NHOL) and to compare the expression with clinicopathological parameters. Study group included 15 cases each of HOL, NHOL and control group included 15 healthy volunteers. Both tissues were immunohistochemically stained for podoplanin (D2-40) antibody. No statistical significant difference was observed between the study and control groups for expression of podoplanin but significant difference was observed on comparison of podoplanin scores between HOL and NHOL. Statistical significant difference was observed when the podoplanin expression in the epithelium and the lymphatic vessel density were correlated with the histologic grading of HOL and NHOL. Expression of podoplanin was greater in NHOL as compared with HOL, this supports the fact that NHOL has a greater risk of malignant transformation when compared with HOL. Podoplanin expression, lymphangiogenesis, and lymphatic vessel density increased with increasing grades of dysplasia, suggesting that cellular modeling and motility is increased as the grade of dysplasia advances. Thus suggesting podoplanin can be used as a prognostic marker to determine the malignant potential in oral leukoplakias.

Utility of Tissue Transglutaminase Immunohistochemistry in Pediatric Duodenal Biopsies: Patterns of Expression and Role in Celiac Disease—A Clinicopathologic Review

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Saeeda Almarzooqi

2013-01-01

Full Text Available Tissue transglutaminase (tTG is a ubiquitous multifunctional protein. It has roles in various cellular processes. tTG is a major target of autoantibodies in celiac disease, and its expression by immunohistochemistry in pediatric celiac disease has not been fully examined. We studied tTG expression in 78 pediatric duodenal biopsies by utilizing an antibody to transglutaminase 2. Serum tTG was positive in all celiac cases evaluated. Serum antiserum endomysial antibody (EMA and tTG were negative in all control subjects and in inflammatory bowel disease and eosinophilic gastroenteritis. There was a statistically significant difference between cases of celiac disease and normal controls in terms of tTG immunohistochemical staining in duodenal biopsies surface epithelium ( value = 0.0012. There was no significant statistical difference in terms of staining of the villous surface or crypt between the cases of celiac disease and cases with IBD ( value = 0.5970 and 0.5227, resp.. There was no detected correlation between serum tTG values and immunohistochemical positivity on duodenal biopsy in cases of celiac disease ( value = 1. There was no relationship between Marsh classification and positivity of villous surface for tTG ( value = 0.4955. We conclude that tTG has limited utility in diagnosis of celiac disease in pediatric duodenal biopsies.

Semi-automatic identification of punching areas for tissue microarray building: the tubular breast cancer pilot study

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Beltrame Francesco

2010-11-01

Full Text Available Abstract Background Tissue MicroArray technology aims to perform immunohistochemical staining on hundreds of different tissue samples simultaneously. It allows faster analysis, considerably reducing costs incurred in staining. A time consuming phase of the methodology is the selection of tissue areas within paraffin blocks: no utilities have been developed for the identification of areas to be punched from the donor block and assembled in the recipient block. Results The presented work supports, in the specific case of a primary subtype of breast cancer (tubular breast cancer, the semi-automatic discrimination and localization between normal and pathological regions within the tissues. The diagnosis is performed by analysing specific morphological features of the sample such as the absence of a double layer of cells around the lumen and the decay of a regular glands-and-lobules structure. These features are analysed using an algorithm which performs the extraction of morphological parameters from images and compares them to experimentally validated threshold values. Results are satisfactory since in most of the cases the automatic diagnosis matches the response of the pathologists. In particular, on a total of 1296 sub-images showing normal and pathological areas of breast specimens, algorithm accuracy, sensitivity and specificity are respectively 89%, 84% and 94%. Conclusions The proposed work is a first attempt to demonstrate that automation in the Tissue MicroArray field is feasible and it can represent an important tool for scientists to cope with this high-throughput technique.

HPASubC: A suite of tools for user subclassification of human protein atlas tissue images.

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Cornish, Toby C; Chakravarti, Aravinda; Kapoor, Ashish; Halushka, Marc K

2015-01-01

The human protein atlas (HPA) is a powerful proteomic tool for visualizing the distribution of protein expression across most human tissues and many common malignancies. The HPA includes immunohistochemically-stained images from tissue microarrays (TMAs) that cover 48 tissue types and 20 common malignancies. The TMA data are used to provide expression information at the tissue, cellular, and occasionally, subcellular level. The HPA also provides subcellular data from confocal immunofluorescence data on three cell lines. Despite the availability of localization data, many unique patterns of cellular and subcellular expression are not documented. To get at this more granular data, we have developed a suite of Python scripts, HPASubC, to aid in subcellular, and cell-type specific classification of HPA images. This method allows the user to download and optimize specific HPA TMA images for review. Then, using a playstation-style video game controller, a trained observer can rapidly step through 10's of 1000's of images to identify patterns of interest. We have successfully used this method to identify 703 endothelial cell (EC) and/or smooth muscle cell (SMCs) specific proteins discovered within 49,200 heart TMA images. This list will assist us in subdividing cardiac gene or protein array data into expression by one of the predominant cell types of the myocardium: Myocytes, SMCs or ECs. The opportunity to further characterize unique staining patterns across a range of human tissues and malignancies will accelerate our understanding of disease processes and point to novel markers for tissue evaluation in surgical pathology.

HPASubC: A suite of tools for user subclassification of human protein atlas tissue images

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Cornish, Toby C.; Chakravarti, Aravinda; Kapoor, Ashish; Halushka, Marc K.

2015-01-01

Background: The human protein atlas (HPA) is a powerful proteomic tool for visualizing the distribution of protein expression across most human tissues and many common malignancies. The HPA includes immunohistochemically-stained images from tissue microarrays (TMAs) that cover 48 tissue types and 20 common malignancies. The TMA data are used to provide expression information at the tissue, cellular, and occasionally, subcellular level. The HPA also provides subcellular data from confocal immunofluorescence data on three cell lines. Despite the availability of localization data, many unique patterns of cellular and subcellular expression are not documented. Materials and Methods: To get at this more granular data, we have developed a suite of Python scripts, HPASubC, to aid in subcellular, and cell-type specific classification of HPA images. This method allows the user to download and optimize specific HPA TMA images for review. Then, using a playstation-style video game controller, a trained observer can rapidly step through 10's of 1000's of images to identify patterns of interest. Results: We have successfully used this method to identify 703 endothelial cell (EC) and/or smooth muscle cell (SMCs) specific proteins discovered within 49,200 heart TMA images. This list will assist us in subdividing cardiac gene or protein array data into expression by one of the predominant cell types of the myocardium: Myocytes, SMCs or ECs. Conclusions: The opportunity to further characterize unique staining patterns across a range of human tissues and malignancies will accelerate our understanding of disease processes and point to novel markers for tissue evaluation in surgical pathology. PMID:26167380

[Pheochromocytoma--pathohistologic and immunohistochemical aspects].

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Tatić, Svetislav; Havelka, Marija; Paunović, Ivan; Bozić, Vesna; Diklic, Aleksandar; Brasanac, Dimitrije; Janković, Radovan; Jancić-Zguricas, Marija

2002-07-01

Pheochromocytoma originates in chromaffin cells of the adrenal medulla. Its incidence is similar in both sexes and most frequent between the ages of thirty and fifty. Multiple and bilateral pheochromocytomas constitute 5 to 10 percent of all cases. Pheochromocytoma occurs sporadically or is related to family syndromes such as: syndrome of multiple endocrine neoplasia--MEN IIA and IIB, neurofibromatosis (von Recklinghausen's disease), von Hippel-Lindau's disease, Sturge-Weber's syndrome, and tuberous sclerosis. Cases in a family usually occur at a younger age and are mostly bilateral and with more aggressive biological behaviour. The aim of the study was to make histomorphological and immunohistochemical analyses of 52 pheochromocytomas. These cases are the surgical material from the Centre of Endocrine Surgery, Institute of Endocrinology, Diabetes, and Metabolic Disorders, Clinical Centre of Serbia, Belgrade, over the period from 1974 to 1997. Frozen and fixed sections, which were cut from paraffinembedded material and stained by both hematoxylin-eosin and PAS, were used in order to make pathohistological diagnoses. The expression of chromogranin A, S-100 protein and ACTH was examined using the PAP method, while neuronspecific enolase (NSE), synaptophysin and neurofilament were examined by the APAAP method with appropriate antibodies (DAKO). The patients were between 4 and 65 years of age (average age 38.5) and there were 28 females (63.64%) and 16 males (36.36%). The largest pheochromocytoma had the diameter of 12 cm, and weight of pheochromocytomas in question was from 13.5 to 370 grams, the average weight being 83.4 grams. On gross examination, the tumours proved to be well-defined, either by fibrous capsule, or by adrenocortical tissue. The cross-sections of tumours were mainly of pale red-grayish colour, and showed numerous foci of necrosis, haemorrhage and cystic softening. Histological appearance of pheochromocytomas was with significant irregularities in

Modified Field's staining--a rapid stain for Trichomonas vaginalis.

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Afzan, M Yusuf; Sivanandam, S; Kumar, G Suresh

2010-10-01

Trichomonas vaginalis, a flagellate protozoan parasite commonly found in the human genitourinary tract, is transmitted primarily by sexual intercourse. Diagnosis is usually by in vitro culture method and staining with Giemsa stain. There are laboratories that use Gram stain as well. We compared the use of modified Field's (MF), Giemsa, and Gram stains on 2 axenic and xenic isolates of T. vaginalis, respectively. Three smears from every sediment of spun cultures of all 4 isolates were stained, respectively, with each of the stains. We showed that MF staining, apart from being a rapid stain (20 s), confers sharper staining contrast, which differentiates the nucleus and the cytoplasm of the organism when compared to Giemsa and Gram staining especially on parasites from spiked urine samples. The alternative staining procedure offers in a diagnostic setting a rapid stain that can easily visualize the parasite with sharp contrasting characteristics between organelles especially the nucleus and cytoplasm. Vacuoles are more clearly visible in parasites stained with MF than when stained with Giemsa. Copyright © 2010 Elsevier Inc. All rights reserved.

Light, electron microscopic and immunohistochemical study of the effect of low-dose aspirin during the proestrus phase on rat endometrium in the preimplantation period.

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Ateş, Utku; Baka, Meral; Turgut, Mehmet; Uyanikgil, Yiğit; Ulker, Sibel; Yilmaz, Ozlem; Tavmergen, Erol; Yurtseven, Mine

2007-04-01

To evaluate structural alterations in rat endometrium at preimplantation following treatment with aspirin beginning from proestrus by light microscopy, electron microscopy and immunohistochemical techniques. Twenty rats were divided into control (n = 10) and experimental (n = 10) groups. Experimental rats were treated with low-dose aspirin daily (2 mg/kg/day) during estrus, beginning from the proestrus phase, mated at end of cycle and treated with aspirin. Untreated pregnant rats were the control group. Rats in both groups were sacrificed at the 84th pregnancy hour; the uterus was rapidly removed and dissected free of surrounding adipose tissue. Uteri specimens from nonpregnant rats were transferred into fixative solution and processed for light, electron microscopic and immunohistochemical study. Light and electron microscopy of endometrium from control rats conformed to mid-diestrus phase; endometrial histology of the aspirin-treated group conformed to late diestrus phase. The endometrial layer was significantly thicker in the aspirin-treated group compared to the untreated control group (p <0.001). No significant difference was found in vessel number between groups. Staining with alphaV integrin was more dense in the aspirin-treated group. Based on histologic findings, we suggest low-dose aspirin has positive effects on preparing endometrium before implantation.

Selfish spermatogonial selection: evidence from an immunohistochemical screen in testes of elderly men.

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Jasmine Lim

Full Text Available The dominant congenital disorders Apert syndrome, achondroplasia and multiple endocrine neoplasia-caused by specific missense mutations in the FGFR2, FGFR3 and RET proteins respectively-represent classical examples of paternal age-effect mutation, a class that arises at particularly high frequencies in the sperm of older men. Previous analyses of DNA from randomly selected cadaveric testes showed that the levels of the corresponding FGFR2, FGFR3 and RET mutations exhibit very uneven spatial distributions, with localised hotspots surrounded by large mutation-negative areas. These studies imply that normal testes are mosaic for clusters of mutant cells: these clusters are predicted to have altered growth and signalling properties leading to their clonal expansion (selfish spermatogonial selection, but DNA extraction eliminates the possibility to study such processes at a tissue level. Using a panel of antibodies optimised for the detection of spermatocytic seminoma, a rare tumour of spermatogonial origin, we demonstrate that putative clonal events are frequent within normal testes of elderly men (mean age: 73.3 yrs and can be classed into two broad categories. We found numerous small (less than 200 cells cellular aggregations with distinct immunohistochemical characteristics, localised to a portion of the seminiferous tubule, which are of uncertain significance. However more infrequently we identified additional regions where entire seminiferous tubules had a circumferentially altered immunohistochemical appearance that extended through multiple serial sections that were physically contiguous (up to 1 mm in length, and exhibited enhanced staining for antibodies both to FGFR3 and a marker of downstream signal activation, pAKT. These findings support the concept that populations of spermatogonia in individual seminiferous tubules in the testes of older men are clonal mosaics with regard to their signalling properties and activation, thus fulfilling

Immunohistochemical expression of CD 10 in Cutaneous basal ,and Squamous Cell Carcinomas

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AIAD, H.A.; HANOUT, H.M.

2007-01-01

Background: CD 10 is a zinc-dependent metallo peptidase known as common acute lymphoblastic leukemia antigen (CALLA). Although CD I 0 expression has been investigated in some cutaneous tumors, to our knowledge, data regarding its expression in cutaneous epithelial neoplasms are very limited. We aimed to determine the immunohistochemical expression of Cd 10 in basal cell carcinoma (BCC) and squamous cell carcinoma (Succ) and to associate it with the available clinico pathological parameters in both tumors. Patients and Methods: This study included 16 Succ and 21 BCC cases (17 solid type, 2 morphea type and 2 adenoid basal types). BCC cases were divided into 12 cases with microscopic infiltrative base and 9 cases with well-circumscribed base. The localization of anti-CD 10 to the tumor and/or stromal cells was determined in each case. Results: Positive CD 10 staining was identified as brown cytoplasmic, with or without cell membrane staining. In all the 16 SCC cases, tumor cells failed to stain with CD 10 in contrast to the stromal cells that showed CD 10 expression in 13 cases (81%). In BCC cases, the expression of CD 10 was noted in tumor cells in 10 cases (476%) and in stromal cells of 20 cases (95.24%). Most of CD 10+ (7/10) BCC showed well-circumscribed deep margin, however, most of CD 10- cases (9/11) showed infiltrating base (p=0.030). BCCs with infiltrating deep margins (12 cases) tended to show CD 10 negative basaloid cells (9/12) and CD 10 positive stromal cells (12/12) (p=0.0003). Conclusion: From our results we suggest that CD 10 might be a useful immunohistochemical marker to differentiate between BCC and SCC. At least, if tumor cells were CD 10 positive, this would favor BCC over SCC. Absence of CD 10 in all the SCC and in infiltrating BCC together with its overexpression in the surrounding stromal cells might confer invasive properties to such tumors. However, its relation to other poor prognostic factors needs larger studies to be confirmed

Meningeal Hemangiopericytomas and Meningomas: a Comparative Immunohistochemical and Genetic Study.

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Trabelsi, Saoussen; Mama, Nadia; Chourabi, Maroua; Mastouri, Maroua Haddaji; Ladib, Mohamed; Popov, Sergey; Burford, Anna; Mokni, Moncef; Tlili, Kalthoum; Krifa, Hedi; Jones, Chris; Yacoubi, Mohamed Tahar; Saad, Ali; Brahim, Dorra H'mida-Ben

2015-01-01

The meningeal hemangiopericytoma (MHPC) is a vascular tumor arising from pericytes. Most intracranial MHPCs resemble meningiomas (MNGs) in their clinical presentation and histological features and may therefore be misdiagnosed, despite important differences in prognosis. We report 8 cases of MHPC and 5 cases of MNG collected from 2007 to 2011 from the Neuro-Surgery and Histopathology departments. All 13 samples were re reviewed by two independent pathologists and investigated by immunohistochemistry (IHC) using mesenchymal, epithelial and neuro-glial markers. Additionally, we screened all tumors for a large panel of chromosomal alterations using multiplex ligation probe amplification (MLPA). Presence of the NAB2-STAT6 fusion gene was inferred by immunohistochemical staining for STAT6. Compared with MNG, MHPCs showed strong VIM (100% of cases), CD99 (62%), bcl-2 (87%), and p16 (75%) staining but only focal positivity with EMA (33%) and NSE (37%). The p21 antibody was positive in 62% of MHPC and less than 1% in all MNGs. MLPA data did not distinguish HPC from MNG, with PTEN loss and ERBB2 gain found in both. By contrast, STAT6 nuclear staining was observed in 3 MHPC cases and was absent from MNG. MNG and MHPC comprise a spectrum of tumors that cannot be easily differentiated based on histopathology. The presence of STAT6 nuclear positivity may however be a useful diagnostic marker.

Surface modification of polycaprolactone scaffolds fabricated via selective laser sintering for cartilage tissue engineering

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Chen, Chih-Hao; Lee, Ming-Yih; Shyu, Victor Bong-Hang; Chen, Yi-Chieh; Chen, Chien-Tzung; Chen, Jyh-Ping

2014-01-01

Surface modified porous polycaprolactone scaffolds fabricated via rapid prototyping techniques were evaluated for cartilage tissue engineering purposes. Polycaprolactone scaffolds manufactured by selective laser sintering (SLS) were surface modified through immersion coating with either gelatin or collagen. Three groups of scaffolds were created and compared for both mechanical and biological properties. Surface modification with collagen or gelatin improved the hydrophilicity, water uptake and mechanical strength of the pristine scaffold. From microscopic observations and biochemical analysis, collagen-modified scaffold was the best for cartilage tissue engineering in terms of cell proliferation and extracellular matrix production. Chondrocytes/collagen-modified scaffold constructs were implanted subdermally in the dorsal spaces of female nude mice. Histological and immunohistochemical staining of the retrieved implants after 8 weeks revealed enhanced cartilage tissue formation. We conclude that collagen surface modification through immersion coating on SLS-manufactured scaffolds is a feasible scaffold for cartilage tissue engineering in craniofacial reconstruction. - Highlights: • Selective laser sintered polycaprolactone scaffolds are prepared. • Scaffolds are surface modified through immersion coating with gelatin or collagen. • Collagen-scaffold is the best for cartilage tissue engineering in vitro. • Chondrocytes/collagen-scaffold reveals enhanced cartilage tissue formation in vivo

Surface modification of polycaprolactone scaffolds fabricated via selective laser sintering for cartilage tissue engineering

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Chen, Chih-Hao [Department of Chemical and Materials Engineering, Chang Gung University, Kweishan, Taoyuan 333, Taiwan, ROC (China); Department of Plastic and Reconstructive Surgery, Chang Gung Memorial Hospital, Craniofacial Research Center, Chang Gung University, Kweishann, Taoyuan 333, Taiwan, ROC (China); Lee, Ming-Yih [Graduate Institute of Medical Mechatronics, Chang Gung University, Kweishan, Taoyuan 333, Taiwan, ROC (China); Shyu, Victor Bong-Hang; Chen, Yi-Chieh; Chen, Chien-Tzung [Department of Plastic and Reconstructive Surgery, Chang Gung Memorial Hospital, Craniofacial Research Center, Chang Gung University, Kweishann, Taoyuan 333, Taiwan, ROC (China); Chen, Jyh-Ping, E-mail: jpchen@mail.cgu.edu.tw [Department of Chemical and Materials Engineering, Chang Gung University, Kweishan, Taoyuan 333, Taiwan, ROC (China); Research Center for Industry of Human Ecology, Chang Gung University of Science and Technology, Kweishan, Taoyuan 333, Taiwan, ROC (China)

2014-07-01

Surface modified porous polycaprolactone scaffolds fabricated via rapid prototyping techniques were evaluated for cartilage tissue engineering purposes. Polycaprolactone scaffolds manufactured by selective laser sintering (SLS) were surface modified through immersion coating with either gelatin or collagen. Three groups of scaffolds were created and compared for both mechanical and biological properties. Surface modification with collagen or gelatin improved the hydrophilicity, water uptake and mechanical strength of the pristine scaffold. From microscopic observations and biochemical analysis, collagen-modified scaffold was the best for cartilage tissue engineering in terms of cell proliferation and extracellular matrix production. Chondrocytes/collagen-modified scaffold constructs were implanted subdermally in the dorsal spaces of female nude mice. Histological and immunohistochemical staining of the retrieved implants after 8 weeks revealed enhanced cartilage tissue formation. We conclude that collagen surface modification through immersion coating on SLS-manufactured scaffolds is a feasible scaffold for cartilage tissue engineering in craniofacial reconstruction. - Highlights: • Selective laser sintered polycaprolactone scaffolds are prepared. • Scaffolds are surface modified through immersion coating with gelatin or collagen. • Collagen-scaffold is the best for cartilage tissue engineering in vitro. • Chondrocytes/collagen-scaffold reveals enhanced cartilage tissue formation in vivo.

A comparison study of histochemical staining of various tissues after ...

African Journals Online (AJOL)

mean =5.28) then Carnoy's (mean = 4.00). For Alcian blue and Perl's Prussian blue, the best staining qualities were obtained by Formalin (mean = 4.76 and 5.64 respectively) followed by Carnoy's (mean = 2.88 and 3.92 respectively).

Are the uterine serous carcinomas underdiagnosed? Histomorphologic and immunohistochemical correlates and clinical follow up in high-grade endometrial carcinomas initially diagnosed as high-grade endometrioid carcinoma.

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Hu, Shaomin; Hinson, Jeff L; Matnani, Rahul; Cibull, Michael L; Karabakhtsian, Rouzan G

2018-02-01

Histologic subclassification of high-grade endometrial carcinomas can sometimes be a diagnostic challenge when based on histomorphology alone. Here we utilized immunohistochemical markers to determine the immunophenotype in histologically ambiguous high-grade endometrial carcinomas that were initially diagnosed as pure or mixed high-grade endometrioid carcinoma, aiming to determine the utility of selected immunohistochemical panel in accurate classification of these distinct tumor types, while correlating these findings with the clinical outcome. A total of 43 high-grade endometrial carcinoma cases initially classified as pure high-grade endometrioid carcinoma (n=32), mixed high-grade endometrioid carcinoma/serous carcinoma (n=9) and mixed high-grade endometrioid carcinoma/clear cell carcinoma (n=2) were retrospectively stained with a panel of immunostains, including antibodies for p53, p16, estrogen receptor, and mammaglobin. Clinical follow-up data were obtained, and stage-to-stage disease outcomes were compared for different tumor types. Based on aberrant staining for p53 and p16, 17/43 (40%) of the high-grade endometrial carcinoma cases initially diagnosed as high-grade endometrioid carcinoma were re-classified as serous carcinoma. All 17 cases showed negative staining for mammaglobin, while estrogen receptor was positive in only 6 (35%) cases. The remaining 26 cases of high-grade endometrioid carcinoma showed wild-type staining for p53 in 25 (96%) cases, patchy staining for p16 in 20 (77%) cases, and were positive for mammaglobin and estrogen receptor in 8 (31%) and 19 (73%) cases, respectively, thus the initial diagnosis of high-grade endometrioid carcinoma was confirmed in these cases. In addition, the patients with re-classified serous carcinoma had advanced clinical stages at diagnosis and poorer overall survival on clinical follow-up compared to that of the remaining 26 high-grade endometrioid carcinoma cases. These results indicate that selected

Evaluation of specific neural marker GAP-43 and TH combined with Masson-trichrome staining for forensic autopsy cases with old myocardial infarction.

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Yu, Tian-Shui; Wang, Xu; Zhang, Hai-Dong; Bai, Ru-Feng; Zhao, Rui; Guan, Da-Wei

2018-01-01

It has been a puzzling forensic task to determine the cause of death as a result of old myocardial infarction (OMI) in the absence of recognizable acute myocardial infarction. Recent studies indicated that the heterogeneous cardiac nerve sprouting and sympathetic hyperinnervation at border zones of the infarcted site played important roles in sudden cardiac death (SCD). So, the present study explored the value of growth associated protein-43 (GAP-43) and tyrosine hydroxylase (TH) as objective and specific neural biomarkers combined with Masson-trichrome staining for forensic autopsy cases. Myocardium of left ventricle of 58 medicolegal autopsy cases, 12 OMI cases, 12 acute/OMI cases, and 34 control cases, were immunostained with anti-GAP-43 and anti-TH antibodies. Immunoreactivity of GAP-43 and TH identified nerve fibers and vascular wall in OMI cases and acute/OMI cases. Specifically, TH-positive nerve fibers were abundant at border zones of the infarcted site. There were a few GAP-43 and TH expressions in the control cases. With Masson-trichrome staining, collagen fibers were blue and cardiac muscle fibers were pink in marked contrast with the surrounding tissue, which improved the location of nerve fibers. Thus, these findings suggest that immunohistochemical detection of GAP-43 and TH combined with Masson-trichrome staining can provide the evidence for the medicolegal expertise of SCD due to OMI, and further demonstrate a close relationship between sympathetic hyperinnervation and SCD.

Detection of abnormalities in the superficial zone of cartilage repaired using a tissue engineered construct derived from synovial stem cells

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W Ando

2012-09-01

Full Text Available The present study investigated the surface structure and mechanical properties of repair cartilage generated from a tissue engineered construct (TEC derived from synovial mesenchymal stem cells at six months post-implantation compared to those of uninjured cartilage. TEC-mediated repair tissue was cartilaginous with Safranin O staining, and had comparable macro-scale compressive properties with uninjured cartilage. However, morphological assessments revealed that the superficial zone of TEC-mediated tissue was more fibrocartilage-like, in contrast to the middle or deep zones that were more hyaline cartilage-like with Safranin O staining. Histological scoring of the TEC-mediated tissue was significantly lower in the superficial zone than in the middle and deep zones. Scanning electron microscopy showed a thick tangential bundle of collagen fibres at the most superficial layer of uninjured cartilage, while no corresponding structure was detected at the surface of TEC-mediated tissue. Immunohistochemical analysis revealed that PRG4 was localised in the superficial area of uninjured cartilage, as well as the TEC-mediated tissue. Friction testing showed that the lubrication properties of the two tissues was similar, however, micro-indentation analysis revealed that the surface stiffness of the TEC-repair tissue was significantly lower than that of uninjured cartilage. Permeability testing indicated that the TEC-mediated tissue exhibited lower water retaining capacity than did uninjured cartilage, specifically at the superficial zone. Thus, TEC-mediated tissue exhibited compromised mechanical properties at the superficial zone, properties which need improvement in the future for maintenance of long term repair cartilage integrity.

Detection of abnormalities in the superficial zone of cartilage repaired using a tissue engineered construct derived from synovial stem cells.

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Ando, Wataru; Fujie, Hiromichi; Moriguchi, Yu; Nansai, Ryosuke; Shimomura, Kazunori; Hart, David A; Yoshikawa, Hideki; Nakamura, Norimasa

2012-09-28

The present study investigated the surface structure and mechanical properties of repair cartilage generated from a tissue engineered construct (TEC) derived from synovial mesenchymal stem cells at six months post-implantation compared to those of uninjured cartilage. TEC-mediated repair tissue was cartilaginous with Safranin O staining, and had comparable macro-scale compressive properties with uninjured cartilage. However, morphological assessments revealed that the superficial zone of TEC-mediated tissue was more fibrocartilage-like, in contrast to the middle or deep zones that were more hyaline cartilage-like with Safranin O staining. Histological scoring of the TEC-mediated tissue was significantly lower in the superficial zone than in the middle and deep zones. Scanning electron microscopy showed a thick tangential bundle of collagen fibres at the most superficial layer of uninjured cartilage, while no corresponding structure was detected at the surface of TEC-mediated tissue. Immunohistochemical analysis revealed that PRG4 was localised in the superficial area of uninjured cartilage, as well as the TEC-mediated tissue. Friction testing showed that the lubrication properties of the two tissues was similar, however, micro-indentation analysis revealed that the surface stiffness of the TEC-repair tissue was significantly lower than that of uninjured cartilage. Permeability testing indicated that the TEC-mediated tissue exhibited lower water retaining capacity than did uninjured cartilage, specifically at the superficial zone. Thus, TEC-mediated tissue exhibited compromised mechanical properties at the superficial zone, properties which need improvement in the future for maintenance of long term repair cartilage integrity.

Fascin and EMMPRIN expression in primary mucinous tumors of ovary: a tissue microarray study.

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Alici, Omer; Kefeli, Mehmet; Yildiz, Levent; Baris, Sancar; Karagoz, Filiz; Kandemir, Bedri

2014-12-01

The aim of this study was to compare the expressions of fascin and EMMPRIN in primary malignant, borderline and benign mucinous ovarian tumors, and to investigate the relationship of these markers with tumor progression and their applicability to differential diagnosis. An immunohistochemical study was performed for fascin and EMMPRIN using the tissue microarray technique. Eighty-one cases were included in the study; there were 37 benign, 25 borderline and 19 malignant primary mucinous ovarian tumors. For each case, a total staining score was determined, consisting of scores for extent of staining and intensity of staining. The cases were allocated to negative, weakly positive and strongly positive staining categories, according to the total staining score. Both of the markers were significantly negative in benign tumors as compared with borderline and malignant tumors. There was no significant difference between borderline and malignant groups for both markers. Sixty-eight percent of malignant tumors were stained positive by fascin, while this rate was 40% for borderline mucinous tumors. All malignant tumors were strongly stained positive for EMMPRIN, while this rate was 92% for borderline mucinous tumors. The rest of the cases stained weakly positive. No significant difference in staining score was found between fascin and EMMPRIN expression. In ovarian primary mucinous tumors, fascin and EMMPRIN may play an important role in tumor progression from benign tumor to carcinoma. In that context, EMMPRIN and fascin expression may have potential application in the differential diagnosis of some diagnostically problematic mucinous ovarian tumors. However, the differential diagnostic applicability of EMMPRIN appears to be more limited than that of fascin due to its wide spectrum of staining in mucinous ovarian tumors. Copyright © 2014 Elsevier GmbH. All rights reserved.

Cytological diagnosis of basal cell carcinoma and actinic keratosis, using Papanicolaou and May-Grünwald-Giemsa stained cutaneous tissue smear.

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Christensen, E; Bofin, A; Gudmundsdóttir, I; Skogvoll, E

2008-10-01

Cytology may become the diagnostic method of choice with the advent of new non-invasive treatments for non-melanoma skin cancer, as the sampling technique for cytology entails little tissue disfiguration. The aim of this study was to compare and evaluate the diagnostic performance of scrape cytology using two different cytological staining techniques, and to evaluate additional touch imprint cytology, with that of histopathology of basal cell carcinoma (BCC) and actinic keratosis (AK). We investigated 50 BCC and 28 AK histologically verified lesions, from 41 and 25 patients, respectively. Two separate skin scrape samples and one touch imprint sample were taken from each lesion. The smears were stained with Papanicolaou (Pap) or May-Grünwald-Giemsa (MGG) stains. All cytological specimens were examined in random order by pathologists without knowledge of the histology. Cytodiagnostic results were compared with the histopathological report. Scrape cytodiagnosis agreed with histopathology in 48 (Pap) and 47 (MGG) of the 50 BCC cases, and in 26 of 28 (Pap) and 21 of 26 (MGG) AK cases, yielding sensitivities of 96%, 94%, 93% and 81%, respectively. No significant difference in sensitivity between the two staining methods was found but a trend towards higher Pap sensitivity for AK was noted (P = 0.10). Touch imprint cytology confirmed histopathology in 38 of the 77 cases of BCC and AK. Cytological diagnosis with either Pap or MGG stain for BCC and AK is reliable, and differentiates well between BCC and AK. Imprint cytology proved to be non-diagnostic in half of the examined cases.

Nucleic Acid Amplification Testing and Sequencing Combined with Acid-Fast Staining in Needle Biopsy Lung Tissues for the Diagnosis of Smear-Negative Pulmonary Tuberculosis.

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Jiang, Faming; Huang, Weiwei; Wang, Ye; Tian, Panwen; Chen, Xuerong; Liang, Zongan

2016-01-01

Smear-negative pulmonary tuberculosis (PTB) is common and difficult to diagnose. In this study, we investigated the diagnostic value of nucleic acid amplification testing and sequencing combined with acid-fast bacteria (AFB) staining of needle biopsy lung tissues for patients with suspected smear-negative PTB. Patients with suspected smear-negative PTB who underwent percutaneous transthoracic needle biopsy between May 1, 2012, and June 30, 2015, were enrolled in this retrospective study. Patients with AFB in sputum smears were excluded. All lung biopsy specimens were fixed in formalin, embedded in paraffin, and subjected to acid-fast staining and tuberculous polymerase chain reaction (TB-PCR). For patients with positive AFB and negative TB-PCR results in lung tissues, probe assays and 16S rRNA sequencing were used for identification of nontuberculous mycobacteria (NTM). The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and diagnostic accuracy of PCR and AFB staining were calculated separately and in combination. Among the 220 eligible patients, 133 were diagnosed with TB (men/women: 76/57; age range: 17-80 years, confirmed TB: 9, probable TB: 124). Forty-eight patients who were diagnosed with other specific diseases were assigned as negative controls, and 39 patients with indeterminate final diagnosis were excluded from statistical analysis. The sensitivity, specificity, PPV, NPV, and accuracy of histological AFB (HAFB) for the diagnosis of smear-negative were 61.7% (82/133), 100% (48/48), 100% (82/82), 48.5% (48/181), and 71.8% (130/181), respectively. The sensitivity, specificity, PPV, and NPV of histological PCR were 89.5% (119/133), 95.8% (46/48), 98.3% (119/121), and 76.7% (46/60), respectively, demonstrating that histological PCR had significantly higher accuracy (91.2% [165/181]) than histological acid-fast staining (71.8% [130/181]), P pulmonary tuberculosis. For patients with positive histological AFB and

Immunohistochemical evidence for an endocrine/paracrine role for ghrelin in the reproductive tissues of sheep

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Brown Yvonne A

2005-10-01

Full Text Available Abstract Background The gut hormone, ghrelin, is involved in the neuroendocrine and metabolic responses to hunger. In monogastric species, circulating ghrelin levels show clear meal-related and body weight-related changes. The pattern of secretion and its role in ruminant species is less clear. Ghrelin acts via growth hormone secretagogue receptors (GHSR-1a to alter food intake, fat utilization, and cellular proliferation. There is also evidence that ghrelin is involved in reproductive function. In the present study we used immunohistochemistry to investigate the presence of ghrelin and GHSR-1a in sheep reproductive tissues. In addition, we examined whether ghrelin and GHSR-1a protein expression is developmentally regulated in the adult and fetal ovine testis, and whether there is an association with markers of cellular proliferation, i.e. stem cell factor (SCF and proliferating cell nuclear antigen (PCNA. Methods Antibodies raised against ghrelin and its functional receptor, GHSR-type 1a, were used in standard immunohistochemical protocols on various reproductive tissues collected from adult and fetal sheep. GHSR-1a mRNA presence was also confirmed by in situ hybridisation. SCF and PCNA immunoexpression was investigated in fetal testicular samples. Adult and fetal testicular immunostaining for ghrelin, GHSR-1a, SCF and PCNA was analysed using computer-aided image analysis. Image analysis data were subjected to one-way ANOVA, with differences in immunostaining between time-points determined by Fisher's least significant difference. Results In adult sheep tissue, ghrelin and GHSR-1a immunostaining was detected in the stomach (abomasum, anterior pituitary gland, testis, ovary, and hypothalamic and hindbrain regions of the brain. In the adult testis, there was a significant effect of season (photoperiod on the level of immunostaining for ghrelin (p Conclusion Evidence is presented for the presence of ghrelin and its receptor in various reproductive

Embedding, serial sectioning and staining of zebrafish embryos using JB-4 resin.

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Sullivan-Brown, Jessica; Bisher, Margaret E; Burdine, Rebecca D

2011-01-01

Histological techniques are critical for observing tissue and cellular morphology. In this paper, we outline our protocol for embedding, serial sectioning, staining and visualizing zebrafish embryos embedded in JB-4 plastic resin-a glycol methacrylate-based medium that results in excellent preservation of tissue morphology. In addition, we describe our procedures for staining plastic sections with toluidine blue or hematoxylin and eosin, and show how to couple these stains with whole-mount RNA in situ hybridization. We also describe how to maintain and visualize immunofluorescence and EGFP signals in JB-4 resin. The protocol we outline-from embryo preparation, embedding, sectioning and staining to visualization-can be accomplished in 3 d. Overall, we reinforce that plastic embedding can provide higher resolution of cellular details and is a valuable tool for cellular and morphological studies in zebrafish.

Evaluation of a menstrual cup to collect shed endometrium for in vitro studies.

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Koks, C A; Dunselman, G A; de Goeij, A F; Arends, J W; Evers, J L

1997-09-01

To evaluate whether a menstrual cup is a suitable instrument to collect antegradely shed endometrium for in vitro studies. A prospective, descriptive, cell biological and immunohistochemical study. Tertiary care university medical center. Nine female volunteers with regular cycles. Menstrual effluent was collected with a menstrual cup. Experience with the menstrual cup was described. Cytospin specimens, frozen sections, and cultures were prepared from the obtained menstrual tissue. The acceptability of the menstrual cup. The presence and viability of endometrial tissue was evaluated using immunohistochemical staining and culture outcome. All women except one described the menstrual cup as acceptable. Menstrual effluent contained single cells, clumps of cells, and glandlike structures. After 5 days of culture, the endometrial tissue appeared to be viable. Immunohistochemistry showed positive staining for vimentin in most cytospin specimens, in all cryostat specimens, and in 10 of 17 cultures. Cytokeratin 18 stained most cytospin specimens, all cryostat specimens, and 10 of 17 cultures. Positive staining for BW495/36 was observed in most cytospin specimens, all cryostat specimens, and 11 of 17 cultures. A menstrual cup in an acceptable instrument to collect antegradely shed menstrual tissue. Menstruum contains viable endometrial tissue that can be used for in vitro studies of endometrium and endometriosis.

Histologic and immunohistochemical evaluation of biocompatibility of castor oil polyurethane polymer with calcium carbonate in equine bone tissue.

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Nóbrega, Fernanda S; Selim, Mariana B; Arana-Chavez, Victor E; Correa, Luciana; Ferreira, Márcio P; Zoppa, André L V

2017-10-01

OBJECTIVE To evaluate the efficacy of castor oil polyurethane polymer with calcium carbonate for use in a unicortical ostectomy on the dorsal surface of the third metacarpal bone of horses. ANIMALS 6 adult horses. PROCEDURES A unicortical ostectomy was created on the dorsal surface of both third metacarpal bones of each horse. Castor bean (Ricinus communis) oil polyurethane polymer with calcium carbonate was implanted into the ostectomy on 1 limb, and the ostectomy of the contralateral limb was left unfilled and served as a control sample. Ostectomy sites were evaluated histologically 120 days later. Biopsy specimens were obtained from the interface of bone and polymer or the interface of bone and newly formed tissue; specimens were processed for histomorphometric evaluation by use of light microscopy, immunohistochemical analysis, histochemical analysis, and transmission electron microscopy. RESULTS Osteoconductive activity of the biomaterial was confirmed by the presence of osteoblasts in the biopsy specimens. Absence of a chronic inflammatory response or foreign body reaction indicated biocompatibility. Expression of osteoblast markers was detected in the newly formed tissue. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that castor oil polyurethane polymer with calcium carbonate could be an acceptable compound for use as a bone substitute in horses with fractures in which bone filling is necessary.

Factors influencing extract of Hibiscus sabdariffa staining of rat testes.

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Bassey, R B; Bakare, A A; Peter, A I; Oremosu, A A; Osinubi, A A

2012-08-01

Some plant extracts can be used in biology and medicine to reveal or identify cellular components and tissues. We investigated the effects of time and concentration on staining of histological sections of rat testes by an acidified extract of Hibiscus sabdariffa. An ethanolic extract of H. sabdariffa was diluted using 1% acetic acid in 70% ethanol to stain histological sections of testes at concentrations of 0.2, 0.1 and 0.05 g/ml for 5, 10, 15, 30, 45 and 60 min. The sections of testes were stained deep red. The staining efficiency of H. sabdariffa was greater at a high concentration and required less time to achieve optimal staining. H. sabdariffa is a strongly basic dye that can be used for various diagnostic purposes. Staining time and concentration must be considered to achieve optimal results.

Immunohistochemical evaluation of proliferation, apoptosis and steroidogenic enzymes in the ovary of rats with polycystic ovary.

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Lombardi, Leonardo Augusto; Simões, Ricardo Santos; Maganhin, Carla Cristina; Baracat, Maria Cândida Pinheiro; Silva-Sasso, Gisela Rodrigues; Florencio-Silva, Rinaldo; Soares, José Maria; Baracat, Edmund Chada

2014-07-01

to evaluate the immunohistochemical expression of proliferative, apoptotic and steroidogenic enzyme markers in the ovaries of rats with polycystic ovary syndrome (PCOS). twenty rats were divided into two groups: GCtrl - estrous phase, and PCOS - with polycystic ovaries. The GCtrl animals were subjected to a lighting period from 7 am to 7 pm, while the animals with PCOS group remained with continuous lighting for 60 days. Subsequently, the animals were anesthetized, the ovaries were removed and fixed in 10% formaldehyde, prior to paraffin embedding. Sections were stained using H.E. or subjected to immunohistochemical methods for the detection of Ki-67, cleaved caspase-3, CYP11A1, CYP17A1 and CYP19A1. The results were analyzed using Student's t-test (p ovaries of rats with PCOS, responsible for the high levels of androgens and estradiol.

3D imaging of hematoxylin and eosin stained thick tissues with a sub-femtoliter resolution by using Cr:forsterite-laser-based nonlinear microscopy (Conference Presentation)

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Kao, Chien-Ting; Wei, Ming-Liang; Liao, Yi-Hua; Sun, Chi-Kuang

2017-02-01

Intraoperative assessment of excision tissues during cancer surgery is clinically important. The assessment is used to be guided by the examination for residual tumor with frozen pathology, while it is time consuming for preparation and is with low accuracy for diagnosis. Recently, reflection confocal microscopy (RCM) and nonlinear microscopy (NLM) were demonstrated to be promising methods for surgical border assessment. Intraoperative RCM imaging may enable detection of residual tumor directly on skin cancers patients during Mohs surgery. The assessment of benign and malignant breast pathologies in fresh surgical specimens was demonstrated by NLM. Without using hematoxylin and eosin (H and E) that are common dyes for histopathological diagnosis, RCM was proposed to image in vivo by using aluminum chloride for nuclear contrast on surgical wounds directly, while NLM was proposed to detect two photon fluorescence nuclear contrast from acrdine orange staining. In this paper, we propose and demonstrate 3D imaging of H and E stained thick tissues with a sub-femtoliter resolution by using Cr:forsterite-laser-based NLM. With a 1260 nm femtosecond Cr:forsterite laser as the excitation source, the hematoxylin will strongly enhance the third-harmonic generation (THG) signals, while eosin will illuminate strong fluorescence under three photon absorption. Compared with previous works, the 1260 nm excitation light provide high penetration and low photodamage to the exercised tissues so that the possibility to perform other follow-up examination will be preserved. The THG and three-photon process provides high nonlinearity so that the super resolution in 3D is now possible. The staining and the contrast of the imaging is also fully compatible with the current clinical standard on frozen pathology thus facilitate the rapid intraoperative assessment of excision tissues. This work is sponsored by National Health Research Institutes and supported by National Taiwan University

Hep par-1: a novel immunohistochemical marker for differentiating hepatocellular carcinoma from metastatic carcinoma

International Nuclear Information System (INIS)

Hanif, R.

2014-01-01

To evaluate the diagnostic utility of Hep par-1 in differentiating hepatocellular carcinoma from metastatic carcinoma taking histopathology as a gold standard. Study Design: Comparative cross-sectional study. Place and Duration of Study: Pathology Department, Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore, from April 2007 to February 2008. Methodology: Hep par-1 immunohistochemical stain was performed on 60 cases of liver carcinoma, 30 cases each of metastatic and hepatocellular carcinoma. Information regarding patient age, gender, sign and symptoms, radiographic findings, histological grade of tumour, and expression of Hep par-1 on hepatocellular and metastatic carcinoma were recorded on proforma sheet. Sensitivity, specificity, positive and negative predictive values, and accuracy of Hep par-1 were calculated using the formulas. Results: Hep par-1 expression was noted in 25 out of 30 cases of hepatocellular carcinoma (83%). Out of 30 cases of metastatic carcinoma, only one case expressed staining in < 5% tumour cells and remaining 29 cases showed no reactivity. The age of the patients with hepatocellular carcinoma ranged from 40 to 76 years with a median age of 60.5 years and 40 - 75 years for metastatic carcinomas with a median age of 57.5 years. Conclusion: Hep par-1 is a reliable immunohistochemical marker for cases of hepatocellular carcinoma (HCC). It can be used along with other markers in morphologically difficult cases when differential diagnosis lies between poorly differentiated HCC and metastatic carcinoma of liver. (author)

Immunohistochemical analysis of Clara cell secretory protein expression in a transgenic model of mouse lung carcinogenesis

International Nuclear Information System (INIS)

Hicks, Sarah M.; Vassallo, Jeffrey D.; Dieter, Matthew Z.; Lewis, Cindy L.; Whiteley, Laurence O.; Fix, Andrew S.; Lehman-McKeeman, Lois D.

2003-01-01

Immunohistochemical methods have been widely used to determine the histogenesis of spontaneous and chemically-induced mouse lung tumors. Typically, antigens for either alveolar Type II cells or bronchiolar epithelial Clara cells are studied. In the present work, the morphological and immunohistochemical phenotype of a transgenic mouse designed to develop lung tumors arising from Clara cells was evaluated. In this model, Clara cell-specific transformation is accomplished by directed expression of the SV40 large T antigen (TAg) under the mouse Clara cell secretory protein (CC10) promoter. In heterozygous mice, early lesions at 1 month of age consisted of hyperplastic bronchiolar epithelial cells. These progressed to adenoma by 2 months as proliferating epithelium extended into adjacent alveolar spaces. By 4 months, a large portion of the lung parenchyma was composed of tumor masses. Expression of constitutive CC10 was diminished in transgenic animals at all time points. Only the occasional cell or segment of the bronchiolar epithelium stained positively for CC10 by immunohistochemistry, and all tumors were found to be uniformly negative for staining. These results were corroborated by Western blotting, where CC10 was readily detectable in whole lung homogenate from nontransgenic animals, but not detected in lung from transgenic animals at any time point. Tumors were also examined for expression of surfactant apoprotein C (SPC), an alveolar Type II cell-specific marker, and found to be uniformly negative for staining. These results indicate that, in this transgenic model, expression of CC10, which is widely used to determine whether lung tumors arise from Clara cells, was reduced and subsequently lost during Clara cell tumor progression

Diagnosing periprosthetic infection: false-positive intraoperative Gram stains.

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Oethinger, Margret; Warner, Debra K; Schindler, Susan A; Kobayashi, Hideo; Bauer, Thomas W

2011-04-01

Intraoperative Gram stains have a reported low sensitivity but high specificity when used to help diagnose periprosthetic infections. In early 2008, we recognized an unexpectedly high frequency of apparent false-positive Gram stains from revision arthroplasties. The purpose of this report is to describe the cause of these false-positive test results. We calculated the sensitivity and specificity of all intraoperative Gram stains submitted from revision arthroplasty cases during a 3-month interval using microbiologic cultures of the same samples as the gold standard. Methods of specimen harvesting, handling, transport, distribution, specimen processing including tissue grinding/macerating, Gram staining, and interpretation were studied. After a test modification, results of specimens were prospectively collected for a second 3-month interval, and the sensitivity and specificity of intraoperative Gram stains were calculated. The retrospective review of 269 Gram stains submitted from revision arthroplasties indicated historic sensitivity and specificity values of 23% and 92%, respectively. Systematic analysis of all steps of the procedure identified Gram-stained but nonviable bacteria in commercial broth reagents used as diluents for maceration of periprosthetic membranes before Gram staining and culture. Polymerase chain reaction and sequencing showed mixed bacterial DNA. Evaluation of 390 specimens after initiating standardized Millipore filtering of diluent fluid revealed a reduced number of positive Gram stains, yielding 9% sensitivity and 99% specificity. Clusters of false-positive Gram stains have been reported in other clinical conditions. They are apparently rare related to diagnosing periprosthetic infections but have severe consequences if used to guide treatment. Even occasional false-positive Gram stains should prompt review of laboratory methods. Our observations implicate dead bacteria in microbiologic reagents as potential sources of false-positive Gram

A novel immunohistochemical sequential multi-labelling and erasing technique enables epitope characterization of bone marrow pericytes in primary myelofibrosis

DEFF Research Database (Denmark)

Madelung, Ann; Bzorek, Michael; Bondo, Henrik

2012-01-01

: In Philadelphia (Ph)-negative chronic myeloproliferative neoplasms, increased microvascular density, bizarre vessel architecture and increased number of pericytes are among the distinct histopathological features. The aim of this study was to characterize bone marrow pericytes in primary myelofibrosis (PMF) using...... a novel multi-labelling immunohistochemical technique. Methods and results: Bone marrow biopsies from a normal donor (n = 1) and patients with PMF (n = 3) were subjected to an immunohistochemical sequential multi-labelling and erasing technique (SE-technique). Antigens of interest in the first and....../or second layer were detected with an immunoperoxidase system and visualized with aminoethylcarbazole. After imaging, erasing and blocking of immunoreagents, the slides were stained with a traditional double immunolabelling procedure. In addition, we applied a Photoshop(®) colour palette, creating a single...

CA 15–3 cell lines and tissue expression in canine mammary cancer and the correlation between serum levels and tumour histological grade

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Manuali Elisabetta

2012-06-01

Full Text Available Abstract Background Mammary tumours are the most common malignancy diagnosed in female dogs and a significant cause of mortality and morbidity in this species. Carbohydrate antigen (CA 15–3 is a mucinous glycoprotein aberrantly over-expressed in human mammary neoplasms and one of the most widely used serum tumour markers in women with breast cancer. The aim of this study was to investigate the antigenic analogies of human and canine CA 15–3 and to assess its expression in canine mammary cancer tissues and cell lines. Immunohistochemical expression of CA 15–3 was evaluated in 7 canine mammary cancer cell lines and 50 malignant mammary tumours. As a positive control, the human breast carcinoma cell line MCF7 and tissue were used. To assess CA 15–3 staining, a semi-quantitative method was applied. To confirm the specificity and cross-reactivity of an anti-human CA 15–3 antibody to canine tissues, an immunoblot analysis was performed. We also investigated serum CA 15–3 activity to establish whether its expression could be assigned to several tumour characteristics to evaluate its potential use as a serum tumour marker in the canine mammary oncology field. Results Immunocytochemical analysis revealed CA 15–3 expression in all examined canine mammary cancer cell lines, whereas its expression was confirmed by immunoblot only in the most invasive cells (CMT-W1, CMT-W1M, CMT-W2 and CMT-W2M. In the tissue, an immunohistochemical staining pattern was observed in 34 (68% of the malignant tumours. A high statistical correlation (p = 0.0019 between serum CA 15–3 levels and the degree of tumour proliferation and differentiation was shown, which indicates that the values of this serum marker increase as the tumour stage progresses. Conclusions The results of this study reveal that CA 15–3 is expressed in both canine mammary tumour cell lines and tissues and that serum levels significantly correlate with the histological grade of the

Stain-free histopathology by programmable supercontinuum pulses

DEFF Research Database (Denmark)

Tu, Haohua; Liu, Yuan; Turchinovich, Dmitry

2016-01-01

The preparation, staining, visualization and interpretation of histological images of tissue is well accepted as the gold standard process for the diagnosis of disease. These methods have a long history of development, and are used ubiquitously in pathology, despite being highly time- and labour-...

Utilization of Cell-Transfer Technique for Molecular Testing on Hematoxylin-Eosin-Stained Sections: A Viable Option for Small Biopsies That Lack Tumor Tissues in Paraffin Block.

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Wu, Howard H; Jovonovich, Stephen M; Randolph, Melissa; Post, Kristin M; Sen, Joyashree D; Curless, Kendra; Cheng, Liang

2016-12-01

- In some instances the standard method of doing molecular testing from formalin-fixed, paraffin-embedded block is not possible because of limited tissue. Tumor cell-enriched cell-transfer technique has been proven useful for performing immunocytochemistry and molecular testing on cytologic smears. - To establish the cell-transfer technique as a viable option for isolating tumor cells from hematoxylin-eosin (H&E)-stained slides. - Molecular testing was performed by using the cell-transfer technique on 97 archived H&E-stained slides from a variety of different tumors. Results were compared to the conventional method of molecular testing. - Polymerase chain reaction-based molecular testing via the cell-transfer technique was successfully performed on 82 of 97 samples (85%). This included 39 of 47 cases for EGFR, 10 of 11 cases for BRAF, and 33 of 39 cases for KRAS mutations. Eighty-one of 82 cell-transfer technique samples (99%) showed agreement with previous standard method results, including 4 mutations and 35 wild-type alleles for EGFR, 4 mutations and 6 wild-type alleles for BRAF, and 11 mutations and 21 wild-type alleles for KRAS. There was only 1 discrepancy: a cell-transfer technique with a false-negative >KRAS result (wild type versus G12C). - Molecular testing performed on H&E-stained sections via cell-transfer technique is useful when tissue from cell blocks and small surgical biopsy samples is exhausted and the only available material for testing is on H&E-stained slides.

Placental heat shock proteins: no immunohistochemical evidence for a differential stress response in preterm labour.

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Divers, M J; Bulmer, J N; Miller, D; Lilford, R J

1995-01-01

The aetiology of idiopathic preterm labour remains obscure. The hypothesis that a stress response induced by low-grade bacterial infection in utero-placental tissues was investigated. Distribution of cognate and inducible isoforms of heat shock proteins (HSP) 70 kD, HSP 60 kD and HSP 90 kD were investigated in an immunohistochemical study of placental and decidual tissues before and after labour at varying gestations. Subjects were pregnant women undergoing singleton delivery after idiopathic preterm labour at less than 34 weeks' gestation (n = 23); spontaneous term labour at 37-42 weeks' gestation (n =24); preterm caesarean sections at less than 34 weeks' gestation for preeclampsia or intrauterine growth retardation (n=14); elective caesarean section at 37-42 weeks' gestation for cephalopelvic disproportion (n = 6). HSP expression was constant throughout the third trimester of pregnancy and did not change following the onset of labour, regardless of gestational age. A stress response in decidual tissues as determined by immunohistochemical analysis is apparently not associated with preterm labour.

Immunohistochemical Analysis of Oral Dysplasia: Diagnostic Assessment by Fascin and Podoplanin Expression

International Nuclear Information System (INIS)

Shimamura, Yumiko; Abe, Takahiro; Nakahira, Mitsuhiko; Yoda, Tetsuya; Murata, Shin-ichi; Sugasawa, Masashi

2011-01-01

The aim of this study was to investigate fascin and podoplanin expression in oral dysplasia and carcinoma in situ (CIS) immunohistochemically, and to evaluate their relationship to histopathological diagnosis based on architectural and cytological features. Fascin and podoplanin expression patterns were analyzed immunohistologically in 26 specimens of oral lesions, including benign disease (hyperplasia, papilloma, and others), intraepithelial neoplasia/borderline disease (dysplasia), and malignant disease (CIS, invasive squamous cell carcinoma). Fascin expression was scored into four original categories, and podoplanin expression was scored into five previously established categories. The relationship between the immunohistochemically determined scores of fascin and podoplanin expression and the architectural and cytological features in the hematoxylin-eosin-stained slides was analyzed statistically. The immunostaining scores for fascin and podoplanin were significantly higher in dysplasia and CIS than in benign disease (p=0.0011, p=0.00036), and they were significantly higher in dysplasia than in benign disease (p=0.0087, p=0.0032). In all cases of invasive SCC, fascin was expressed mainly in the cytoplasm of the tumor cells and fascin expression extended from the destruction of the basal layer of the epithelium to the upper layer of the epithelium and podoplanin was expressed in the cytoplasm and membrane of the tumor cells. This was the first report of up-regulation of fascin in oral dysplasia. Our results suggest that it would be helpful for improving the diagnostic accuracy of oral dysplasia and CIS to assess the expression of fascin and podoplanin immunohistochemically

Biochemical and immunohistochemical characterization of proteins in Hürthle cell carcinoma.

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De Keyser, L; Layfield, L; Van Herle, A; Costin, A; Lewin, K

1984-10-01

The present study reports the biochemical and immunohistochemical findings in the cytosol of a Hürthle cell carcinoma as compared with that of normal thyroid tissue. Sephadex G-200 chromatography of the extract derived from a Hürthle cell carcinoma and from normal thyroid tissue revealed three identical pools. Pool I consisted mainly of thyroglobulin (Tg), pool II corresponded to albumin, while pool III contained unidentified low molecular weight fragments which could not be studied further. Hürthle cell carcinoma, pool I, had a Tg content of 12.9 micrograms Tg/mg equivalent tissue and a 127I content of 5,6 mole/mole of Tg. Its sialic acid content was undetectable, however. In pool I of the normal thyroid gland, the respective values were 62.8 micrograms Tg/mg equivalent tissue, 21.3 +/mole 127I/mole Tg, and 15.4 mole sialic acid/mole Tg. The albumin contained in both pools II was shown to be ioidinated at the following levels: 0.025 mole 127I/mole albumin in Hürthle tumor pool II vs 1.28 mole 127I/mole albumin in normal thyroid pool II. Immunohistochemical studies confirmed the presence of Tg and albumin in the malignant Hürthle cells and acini and colloid. Thus, Hürthle cell carcinoma contained Tg and albumin. The Tg content was five times less compared with control tissue. Both proteins (Tg and albumin) were poorly iodinated in Hürthle carcinoma tissue, and the iodination of albumina seemed to be more severely impaired. The site of synthesis of both proteins could not be derived from the present studies.

Objective Quantification of Immune Cell Infiltrates and Epidermal Proliferation in Psoriatic Skin

DEFF Research Database (Denmark)

Soendergaard, Christoffer; Nielsen, Ole H; Skak, Kresten

2015-01-01

assessments by pathologists with the interobserver and intraobserver variation this includes. Automated quantitative assessment of immunohistochemical staining has the potential to objectively extract numerical measures from cell and tissue structures, and allows efficient high throughput analysis in clinical...... research. Published data of manual cell counts in psoriatic skin samples were in this study reevaluated using the digital image analysis (DIA) software. Whole slides immunohistochemically stained for CD3, CD4, CD8, CD45R0, and Ki-67 were scanned and quantitatively evaluated using simple threshold analysis...

HPASubC: A suite of tools for user subclassification of human protein atlas tissue images

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Toby C Cornish

2015-01-01

Full Text Available Background: The human protein atlas (HPA is a powerful proteomic tool for visualizing the distribution of protein expression across most human tissues and many common malignancies. The HPA includes immunohistochemically-stained images from tissue microarrays (TMAs that cover 48 tissue types and 20 common malignancies. The TMA data are used to provide expression information at the tissue, cellular, and occasionally, subcellular level. The HPA also provides subcellular data from confocal immunofluorescence data on three cell lines. Despite the availability of localization data, many unique patterns of cellular and subcellular expression are not documented. Materials and Methods: To get at this more granular data, we have developed a suite of Python scripts, HPASubC, to aid in subcellular, and cell-type specific classification of HPA images. This method allows the user to download and optimize specific HPA TMA images for review. Then, using a playstation-style video game controller, a trained observer can rapidly step through 10′s of 1000′s of images to identify patterns of interest. Results: We have successfully used this method to identify 703 endothelial cell (EC and/or smooth muscle cell (SMCs specific proteins discovered within 49,200 heart TMA images. This list will assist us in subdividing cardiac gene or protein array data into expression by one of the predominant cell types of the myocardium: Myocytes, SMCs or ECs. Conclusions: The opportunity to further characterize unique staining patterns across a range of human tissues and malignancies will accelerate our understanding of disease processes and point to novel markers for tissue evaluation in surgical pathology.

Effects of Melatonin on the Cerebellum of Infant Rat Following Kaolin-Induced Hydrocephalus: a Histochemical and Immunohistochemical Study.

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Uyanıkgil, Yiğit; Turgut, Mehmet; Baka, Meral

2017-02-01

Hydrocephalus is a developmental disorder causing abnormally collected cerebrospinal fluid within the cerebral ventricles. It leads to bigger skulls and many dysfunctions related to the nervous system. Here, we addressed whether exogenous melatonin administration could reverse the clinical features of kaolin-induced hydrocephalus in infantile rats. A controlled double-blinded study was conducted in 2-week-old 45 Wistar albino rats, which were divided into three groups: Group A, the control group, received intracisternal sham injection with solely the needle insertion; group B, the hydrocephalus group, was treated with isotonic NaCl after kaolin injection; and group C, the hydrocephalus + melatonin group, was given i.p. exogenous melatonin at a dose of 0.5 mg/100 g body weight after kaolin injection. Histological and immunohistochemical analyses were performed after the induction of hydrocephalus and melatonin administration. Glial fibrillary acidic protein was stained by immunohistochemical method. TUNEL method was used to define and quantitate apoptosis in the cerebellar tissues. Statistical analysis was performed by nonparametric Kruskal-Wallis H test, and once significance was determined among means, post hoc pairwise comparisons were carried out using Mann-Whitney U test. We found that melatonin administration significantly ameliorated ratio of substantia grisea area/substantia alba area in the cerebellum of infantile rats. Histologically, there was a significant reduction in the number of cerebellar apoptotic cells after the hydrocephalus induced by kaolin (P cerebellum were reversed by systemic melatonin administration in infantile rats with kaolin-induced hydrocephalus. Nevertheless, further studies are needed to suggest melatonin as a candidate protective drug in children with hydrocephalus.

Staining Against Phospho-H2AX (gamma-H2AX) as a Marker for DNA Damage and Genomic Instability in Cancer Tissues and Cells

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Nagelkerke, A.P.; Span, P.N.

2016-01-01

Phospho-H2AX or gamma-H2AX- is a marker of DNA double-stranded breaks and can therefore be used to monitor DNA repair after, for example, irradiation. In addition, positive staining for phospho-H2AX may indicate genomic instability and telomere dysfunction in tumour cells and tissues. Here, we

Basement membrane changes in breast cancer detected by immunohistochemical staining for laminin

DEFF Research Database (Denmark)

Albrechtsen, R; Nielsen, M; Wewer, U

1981-01-01

The distribution of the basement membrane glycoprotein laminin was studied by the immunoperoxidase technique in benign and malignant human breast tissue and in axillary lymph nodes from patients with breast cancer. An antiserum prepared against rat laminin was used. The specificity...

Mercury localization in mouse kidney over time: autoradiography versus silver staining

International Nuclear Information System (INIS)

Rodier, P.M.; Kates, B.; Simons, R.

1988-01-01

Several methods of silver staining have been employed to localize mercury in tissue, under the assumption that the techniques represent total Hg, but recent reports have suggested that these stains are specific for a limited fraction of the Hg present in some samples. Magos et al. hypothesized that the stains actually vary with inorganic mercury content. The purpose of the present study was to compare localization by radiolabeling to localization by one silver stain, the photoemulsion histochemical technique, in tissues prepared to contain a range of levels of total Hg and a range of levels of inorganic Hg. Mice dosed with 8 mg Hg/kg as MeHg were killed 24 hr, 1 week, or 2 weeks after exposure, to allow a decrease in total Hg and an increase in the proportion of demethylated Hg over time. Mice dosed with 4 mg Hg/kg as HgCl 2 provided samples in which all the Hg present was in the inorganic form. Atomic absorption of kidneys of mice dosed with MeHg showed that total Hg fell from 55 micrograms/g to 39 to 25 over 2 weeks, while the inorganic fraction climbed from about 2 to 27 to 35%. Grain counts from autoradiographs of 203 Hg-labeled sections correlated with total Hg content at +0.88, but silver staining was correlated with inorganic Hg content, appearing only at late termination times in MeHg-exposed animals, but soon after dosing in mice exposed to inorganic Hg. The photoemulsion histochemical technique revealed a substance strictly localized in the proximal tubules, while autoradiographs and grain counts showed total Hg to be present throughout the kidney tissue. These results support the contention that silver stains are selective for inorganic Hg

Merkel Cell Carcinoma with Spontaneous Regression: A Case Report and Immunohistochemical Study

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Hitoshi Terui

2016-02-01

Full Text Available Merkel cell carcinoma (MCC is an aggressive neuroendocrine carcinoma that only rarely regresses spontaneously. Since little is known about the immunological mechanisms involved in the spontaneous regression of MCC, we describe a case of MCC with spontaneous regression and employed immunohistochemical staining for cytotoxic and immunosuppressive molecules to investigate possible mechanisms involved in the spontaneous regression of MCC. Interestingly, compared to conventional MCC, tumor-infiltrating lymphocytes in MCC with spontaneous regression contained higher numbers of CD8+ cells and granulysin-bearing cells and lower numbers of CD206+ cells. Our present study suggests one of the possible reasons for the spontaneous regression of MCC.

[Study of methods of decalcification for making united slices of tooth and affiliated periodontic tissues].

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Wang, Yu; Mu, Ya-bing; Miao, Lei-ying; Sun, Hong-chen; Li, Cheng-ku

2007-03-01

To study the methods of decalcification for making united slices of tooth and affiliated periodontic tissues. Twenty-one samples containing dog molars and affiliated periodontic tissues were divided into seven mean groups. The pH value of solution, time of decalcification, weight and volume of samples, and content of decalcified calcium were detected. The slices were observed by HE, specific, and immunohistochemical stain. The velocity of decalcification increased with decrease of solution pH. The weight of samples lightened by 37.61%, the volume reduced by 25.97% on average, and calcium decalcified was 174.49 mg per gram humid samples. The EDTA decalcification was slowest, but it was best. Decalcification was fast in Plank-Rycho solution while the section was worst, and faster in the formyl solution containing aluminium chloride than in EDTA, and the section was better. The 50% formyl solution containing aluminium chloride is an ideal decalcifying solution.

Lectins stain cells differentially in the coral, Montipora capitata

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Work, Thierry M.; Farah, Yael

2014-01-01

A limitation in our understanding of coral disease pathology and cellular pathogenesis is a lack of reagents to characterize coral cells. We evaluated the utility of plant lectins to stain tissues of a dominant coral, Montipora capitata, from Hawaii. Of 22 lectins evaluated, nine of these stained structures in the upper or basal body wall of corals. Specific structures revealed by lectins that were not considered distinct or evident on routine hematoxylin and eosin sections of coral tissues included apical and basal granules in gastrodermis and epidermis, cnidoglandular tract and actinopharynx cell surface membranes, capsules of mature holotrichous isorhizas, and perivitelline and periseminal cells. Plant lectins could prove useful to further our understanding of coral physiology, anatomy, cell biology, and disease pathogenesis.

Preparation of colloidal gold for staining proteins electrotransferred onto nitrocellulose membranes.

Science.gov (United States)

Yamaguchi, K; Asakawa, H

1988-07-01

This paper describes a simple method of preparing colloidal gold for staining protein blots. Colloidal gold was prepared from 0.005 or 0.01% HAuCl4 by the addition of formalin as a reductant and potassium hydroxide. Staining of small cell carcinoma tissue extract blotted onto nitrocellulose membranes with this colloidal gold solution resulted in the appearance of a large number of clear wine-red bands. The sensitivity of gold staining was 60 times higher than that of Coomassie brilliant blue staining and almost comparable to that of silver staining of proteins in polyacrylamide gel. The sensitivity of this method was also satisfactory in comparison with that of enzyme immunoblotting. The colloidal gold prepared by this method is usable for routine work.

Analysis of human papilloma virus in oral squamous cell carcinoma using p16: An immunohistochemical study

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Patil, S.; Rao, R. S.; Amrutha, N.; Sanketh, D. S.

2014-01-01

Aims: The aim of this study is to evaluate the expression of human papilloma virus (HPV) in oral squamous cell carcinoma (OSCC) and to correlate the association of HPV in histological grades of OSCC using p16 (p16INK4a) immunohistochemistry (IHC). Subjects and Methods: This study consists of 30 histological diagnosed cases of OSCC (10-well-differentiated oral squamous cell carcinoma [WDOSCC], 10-moderately differentiated oral squamous cell carcinoma [MDOSCC] and 10-poorly differentiated oral squamous cell carcinoma [PDOSCC]). The sections were subjected to IHC procedure using p16. Two parameters in immunohistochemical p16 expression were evaluated by 3 observers based on the criteria by Galgano M. Tetal (2010) (a) percentage of p16 positive cases (b) pattern of p16 staining in various grades of OSCC. Statistical Analysis Used: Kappa test. Results: Totally, 30 samples of 0SCC, p16 positivity was noted in 26/30 (86.66%). Of 26 positive cases, p16 staining was positive in 7/10 (70%) of WDOSCC, 9/10 (90%) in MDOSCC and, 10/10 (100%) PDOSCC. Incidentally, we also found single dispersed cell staining in WDOSCC, patchy staining in MDOSCC and more diffuse staining pattern predominant in PDOSCC. Conclusions: Our study revealed an association between HPV and OSCC. Diffuse staining pattern was noted in PDOSCC, which in turn depicts the increase viral overload, which might have an influence on its aggressive behavior. PMID:24818098

Proteomics analysis of apoptosis-regulating proteins in tissues with different radiosensitivity

International Nuclear Information System (INIS)

An, Jeung-Hee; Seong, Jin-Sil

2006-01-01

The aim of this study was to identify of radiosusceptibility proteins in tissues with different radiosensitivity. C3H/HeJ mice were exposed to 10 Gy. The tissues were processed for proteins extraction and were analyzed by 2-dimensional electrophoresis. The proteins were identified by matrix-assisted laser desorption ionizing time-of-flight mass spectrometry and validated by immunohistochemical staining and Western blotting. The peaks of apoptosis levels were 35.3±1.7% and 0.6±0.2% in the spleen and the liver, respectively, after ionizing radiation. Analysis of liver tissue showed that the expression level of reactive oxygen species (ROS) related proteins such as cytochrome c, glutathione S transferase, NADH dehydrogenase and peroxiredoxin VI increased after radiation. The expression level of cytochrome c increased to 3-fold after ionizing radiation in both tissues. However in spleen tissue, the expression level of various kinds of apoptosis regulating proteins increased after radiation. These involved iodothyronine, CD 59A glycoprotein precursor, fas antigen and tumor necrosis factor -inducible protein TSG-6nprecursor after radiation. The difference in the apoptosis index between the liver and spleen tissues is closely associated with the expression of various kinds of apoptosis-related proteins. The result suggests that the expression of apoptosis-related protein and redox proteins play important roles in this radiosusceptibility. (author)

Matrix metalloproteases and tissue inhibitors of metalloproteinases in medial plica and pannus-like tissue contribute to knee osteoarthritis progression.

Science.gov (United States)

Yang, Chih-Chang; Lin, Cheng-Yu; Wang, Hwai-Shi; Lyu, Shaw-Ruey

2013-01-01

Osteoarthritis (OA) is characterized by degradation of the cartilage matrix, leading to pathologic changes in the joints. However, the pathogenic effects of synovial tissue inflammation on OA knees are not clear. To investigate whether the inflammation caused by the medial plica is involved in the pathogenesis of osteoarthritis, we examined the expression of matrix metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs), interleukin (IL)-1β, and tumor necrosis factor (TNF)-α in the medial plica and pannus-like tissue in the knees of patients with medial compartment OA who underwent either arthroscopic medial release (stage II; 15 knee joints from 15 patients) or total knee replacement (stage IV; 18 knee joints from 18 patients). MMP-2, MMP-3, MMP-9, IL-1β, and TNF-α mRNA and protein levels measured, respectively, by quantitative real-time PCR and Quantibody human MMP arrays, were highly expressed in extracts of medial plica and pannus-like tissue from stage IV knee joints. Immunohistochemical staining also demonstrated high expression of MMP-2, MMP-3, and MMP-9 in plica and pannus-like tissue of stage IV OA knees and not in normal cartilage. Some TIMP/MMP ratios decreased significantly in both medial plica and pannus-like tissue as disease progressed from stage II to stage IV. Furthermore, the migration of cells from the pannus-like tissue was enhanced by IL-1β, while plica cell migration was enhanced by TNF-α. The results suggest that medial plica and pannus-like tissue may be involved in the process of cartilage degradation in medial compartment OA of the knee.

Matrix metalloproteases and tissue inhibitors of metalloproteinases in medial plica and pannus-like tissue contribute to knee osteoarthritis progression.

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Chih-Chang Yang

Full Text Available Osteoarthritis (OA is characterized by degradation of the cartilage matrix, leading to pathologic changes in the joints. However, the pathogenic effects of synovial tissue inflammation on OA knees are not clear. To investigate whether the inflammation caused by the medial plica is involved in the pathogenesis of osteoarthritis, we examined the expression of matrix metalloproteinases (MMPs, tissue inhibitors of metalloproteinases (TIMPs, interleukin (IL-1β, and tumor necrosis factor (TNF-α in the medial plica and pannus-like tissue in the knees of patients with medial compartment OA who underwent either arthroscopic medial release (stage II; 15 knee joints from 15 patients or total knee replacement (stage IV; 18 knee joints from 18 patients. MMP-2, MMP-3, MMP-9, IL-1β, and TNF-α mRNA and protein levels measured, respectively, by quantitative real-time PCR and Quantibody human MMP arrays, were highly expressed in extracts of medial plica and pannus-like tissue from stage IV knee joints. Immunohistochemical staining also demonstrated high expression of MMP-2, MMP-3, and MMP-9 in plica and pannus-like tissue of stage IV OA knees and not in normal cartilage. Some TIMP/MMP ratios decreased significantly in both medial plica and pannus-like tissue as disease progressed from stage II to stage IV. Furthermore, the migration of cells from the pannus-like tissue was enhanced by IL-1β, while plica cell migration was enhanced by TNF-α. The results suggest that medial plica and pannus-like tissue may be involved in the process of cartilage degradation in medial compartment OA of the knee.

Immunohistochemical localization of steroid receptor coactivators in chondrosarcoma: an in vivo tissue microarray study.

Science.gov (United States)

Li, Wei; Fu, Jingshu; Bian, Chen; Zhang, Jiqiang; Xie, Zhao

2014-12-01

Chondrosarcoma is the second most common type of primary bone malignancy following up osteosarcoma, characterized by resistance to conventional chemotherapeutic agents and radiation regimens. The p160 family members steroid receptor coactivator-1 and -3 (SRC-1 and SRC-3) have been implied in the regulation of cancer growth, migration, invasion, metastasis and chemotherapeutic resistance; but we still lack detailed information about the levels of SRCs in chondrosarcoma. In this study, expression of SRC-1 and SRC-3 in chondrosarcoma was examined by immunohistochemistry with tissue microarrays; the four score system (0, 1, 2 and 3) was used to evaluate the staining. The results showed that there were no gender-, site- or age-differences regarding the expression of SRC-1 or SRC-3 (p>0.05); organ (bone or cartilage) -differences were only detected for SRC-1 but not SRC-3 (pchondrosarcoma, may be novel targets for the prognosis and/or treatment of chondrosarcoma, would have opened a new avenue and established foundation for studying chondrosarcoma. Copyright © 2014 Elsevier GmbH. All rights reserved.

Immunohistochemical analysis of the distribution of desmoglein 1 and 2 in the skin of dogs and cats.

Science.gov (United States)

Miragliotta, Vincenzo; Coli, Alessandra; Ricciardi, Maria P; Podestà, Adriano; Abramo, Francesca

2005-11-01

To compare the distribution of desmoglein (Dsg) 1 and 2 in skin specimens obtained from dogs and cats to provide information about the possible role of the density of Dsg 1 and 2 in the localization of lesions attributable to pemphigus foliaceus in these 2 species. Skin biopsy specimens obtained from 4 dogs and 4 cats. Biopsy specimens were collected from the muzzle, bridge of the nose, ear, dorsum, abdomen, area adjacent to the teats, and footpads of each animal. Immunohistochemical analysis was performed on formalin-fixed, paraffin-embedded skin samples by use of a biotinylated mouse monoclonal anti-Dsg 1 and 2 antibody raised against bovine muzzle. Color development was performed by use of the streptavidin-biotin-peroxidase method with a chromogenic substrate. Immunohistochemical staining yielded a positive reaction in skin samples obtained from all anatomic sites. The intensity and distribution of staining were related to the number of layers of the stratum spinosum. No differences were detected between samples obtained from dogs and cats. No differences in intensity of Dsg 1 and 2 antigen were observed in the stratum spinosum between skin samples obtained from dogs and cats. Analysis of this result suggests that factors other than the distribution of Dsg may be responsible for the differences in localization of primary clinical lesions in dogs and cats with pemphigus foliaceus.

Modified Cajal's trichrome stain as a diagnostic aid in the study of epithelial pathology

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Karpagaselvi Sanjai

2017-01-01

Full Text Available Background: Diagnosis of initial epithelial pathology maybe difficult in Squamous Cell Carcinoma (SCC, Carcinoma In Situ and other atypical epithelial malignancies, under routine Haematoxylin and Eosin (H and E stain. The detection of minor basement membrane alterations in doubtful cases is both time consuming and confusing. Aims: To evaluate efficacy of Modified Cajal's Trichrome Stain (CTS in relation to Haematoxylin and Eosin for study of epithelial dysplasia, carcinoma in situ, micro invasive SCC, frank SCC, and SCC in lymph nodes. Materials and Methods: Formalin-fixed, paraffin-embedded tissue blocks of mild epithelial dysplasia (n = 2, moderate epithelial dysplasia (n = 2, severe epithelial dysplasia (n = 4, carcinoma in situ (n = 1, micro-invasive SCC (n = 4, verrucous carcinoma (n = 1, and frank OSCC (n = 5 were stained with CTS and H&E. The sections were compared based on set histopathological criteria. Results and Conclusion: In SCC cases stained with CTS, invasion into connective tissue and keratin pearls were strikingly evident. Depth of invasion could be more accurately determined. Tumour cells in lymph node were intensely contrasted and easily discernible. Thus, CTS is a good differential stain, clearly delineating the epithelial elements from the connective tissue elements visually. This helps in tracing the basement membrane very clearly. It is an economic, rapid and easy to use method which cannot replace Haematoxylin and Eosin stain in cancer diagnosis, but can definitely be used adjunctive to it. Prompt diagnosis is crucial to effective treatment, and this stain assists in early and rapid diagnosis of cancer.

Connective tissue growth factor stimulates the proliferation, migration and differentiation of lung fibroblasts during paraquat-induced pulmonary fibrosis.

Science.gov (United States)

Yang, Zhizhou; Sun, Zhaorui; Liu, Hongmei; Ren, Yi; Shao, Danbing; Zhang, Wei; Lin, Jinfeng; Wolfram, Joy; Wang, Feng; Nie, Shinan

2015-07-01

It is well established that paraquat (PQ) poisoning can cause severe lung injury during the early stages of exposure, finally leading to irreversible pulmonary fibrosis. Connective tissue growth factor (CTGF) is an essential growth factor that is involved in tissue repair and pulmonary fibrogenesis. In the present study, the role of CTGF was examined in a rat model of pulmonary fibrosis induced by PQ poisoning. Histological examination revealed interstitial edema and extensive cellular thickening of interalveolar septa at the early stages of poisoning. At 2 weeks after PQ administration, lung tissue sections exhibited a marked thickening of the alveolar walls with an accumulation of interstitial cells with a fibroblastic appearance. Masson's trichrome staining revealed a patchy distribution of collagen deposition, indicating pulmonary fibrogenesis. Western blot analysis and immunohistochemical staining of tissue samples demonstrated that CTGF expression was significantly upregulated in the PQ-treated group. Similarly, PQ treatment of MRC-5 human lung fibroblast cells caused an increase in CTGF in a dose-dependent manner. Furthermore, the addition of CTGF to MRC-5 cells triggered cellular proliferation and migration. In addition, CTGF induced the differentiation of fibroblasts to myofibroblasts, as was evident from increased expression of α-smooth muscle actin (α-SMA) and collagen. These findings demonstrate that PQ causes increased CTGF expression, which triggers proliferation, migration and differentiation of lung fibroblasts. Therefore, CTGF may be important in PQ-induced pulmonary fibrogenesis, rendering this growth factor a potential pharmacological target for reducing lung injury.

Clinical and immunohistochemical comparative study of the efficacy of carboxytherapy vs platelet-rich plasma in treatment of stretch marks.

Science.gov (United States)

Hodeib, Abeer A; Hassan, Ghada F R; Ragab, Marwa N M; Hasby, Eiman A

2018-01-07

Striae distensae are dermal scars with a linear atrophic depression. The exact origin of striae distensae remains unrevealed, but low expression of collagen and fibronectin genes in the affected tissue was found. Several treatment modalities have been proposed, yet no consistent modality is available. To evaluate and compare the efficacy and safety of carboxytherapy vs platelet-rich plasma (PRP) in treatment of stretch marks. This study included 20 patients with striae alba. Every patient received treatment in the form of PRP injection in their right side (group A) and carboxytherapy session in their left side (group B) every 3-4 weeks for 4 sessions. Skin biopsies were taken before and after treatment, and they were subjected to fibronectin immunohistochemical stain. There was a significant improvement in striae alba in both groups after than before treatment. There was no significant difference between both groups as regards either percentage of improvement, response (grading scale), or patient satisfaction. The fibronectin-stained area was significantly higher in both groups after than before treatment, and it was significantly higher after treatment in group (B) than group (A). Both methods were safe and effective with minimal side effects. There was no significant difference between both methods of treatments. This was confirmed histopathologically by fibronectin expression which is found to be low in striae and increased significantly after treatment. But fibronectin expression was higher in group (B) than (A). © 2018 Wiley Periodicals, Inc.

[Esophageal sarcomatoid carcinoma: report of a case with morphological, immunohistochemical and molecular study].

Science.gov (United States)

Regragui, Asmaa; Lakhdar, Hind; Abderrahman Alaoui Belabbas, Moulay; Amrani, Meryem; Gamra, Lamia; Alaoui Belabbas, Mohamed

2004-05-01

Sarcomatoïd carcinoma is a rare tumor of the esophagus, characterized macroscopically by a polypoid aspect and histologically by the association of spindle cell carcinoma with sarcomatous pleomorphic component. We report here a case of esophagus sarcomatoïd carcinoma. Diagnosis was based on immunohistochemical analysis of tIssue samples. Human papillomavirus (HVP) detection by PCR amplification of DNA extracted from tumoral tIssue was negative, ruling out the role of HPV infection in this tumor.

Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids.

Science.gov (United States)

Farkona, Sofia; Soosaipillai, Antoninus; Filippou, Panagiota; Korbakis, Dimitrios; Serra, Stefano; Rückert, Felix; Diamandis, Eleftherios P; Blasutig, Ivan M

2017-12-01

CUB and zona pellucida-like domain-containing protein 1 (CUZD1) was identified as a pancreas-specific protein and was proposed as a candidate biomarker for pancreatic related disorders. CUZD1 protein levels in tissues and biological fluids have not been extensively examined. The purpose of the present study was to generate specific antibodies targeting CUZD1 to assess CUZD1 expression within tissues and biological fluids. Mouse monoclonal antibodies against CUZD1 were generated and used to perform immunohistochemical analyses and to develop a sensitive and specific enzyme-linked immunosorbent assay (ELISA). CUZD1 protein expression was assessed in various human tissue extracts and biological fluids and in gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant protein. Immunohistochemical staining of CUZD1 in pancreatic tissue showed that the protein is localized to the acinar cells and the lumen of the acini. Western blot analysis detected the protein in pancreatic tissue extract and pancreatic juice. The newly developed ELISA measured CUZD1 in high levels in pancreas and in much lower but detectable levels in several other tissues. In the biological fluids tested, CUZD1 expression was detected exclusively in pancreatic juice. The analysis of gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant CUZD1 suggested that the protein exists in high molecular weight protein complexes. This study describes the development of tools targeting CUZD1 protein, its tissue expression pattern and levels in several biological fluids. These new tools will facilitate future investigations aiming to delineate the role of CUZD1 in physiology and pathobiology. Copyright © 2017 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

CDX2 downregulation is associated with poor differentiation and MMR deficiency in colon cancer

DEFF Research Database (Denmark)

Olsen, J.; Eiholm, Susanne; Kirkeby, L T

2016-01-01

adjacent tissue were fixed in liquid nitrogen for RNA extraction or in formalin and paraffin embedded (FFPE) for immunohistochemical staining. CDX2 mRNA expression was evaluated by RT-qPCR. FFPE sections were stained for MLH1, MSH2, MSH6, PMS2, and CDX2. RESULTS: A total of 191 patient samples were...

Zooming in: high resolution 3D reconstruction of differently stained histological whole slide images

Science.gov (United States)

Lotz, Johannes; Berger, Judith; Müller, Benedikt; Breuhahn, Kai; Grabe, Niels; Heldmann, Stefan; Homeyer, André; Lahrmann, Bernd; Laue, Hendrik; Olesch, Janine; Schwier, Michael; Sedlaczek, Oliver; Warth, Arne

2014-03-01

Much insight into metabolic interactions, tissue growth, and tissue organization can be gained by analyzing differently stained histological serial sections. One opportunity unavailable to classic histology is three-dimensional (3D) examination and computer aided analysis of tissue samples. In this case, registration is needed to reestablish spatial correspondence between adjacent slides that is lost during the sectioning process. Furthermore, the sectioning introduces various distortions like cuts, folding, tearing, and local deformations to the tissue, which need to be corrected in order to exploit the additional information arising from the analysis of neighboring slide images. In this paper we present a novel image registration based method for reconstructing a 3D tissue block implementing a zooming strategy around a user-defined point of interest. We efficiently align consecutive slides at increasingly fine resolution up to cell level. We use a two-step approach, where after a macroscopic, coarse alignment of the slides as preprocessing, a nonlinear, elastic registration is performed to correct local, non-uniform deformations. Being driven by the optimization of the normalized gradient field (NGF) distance measure, our method is suitable for differently stained and thus multi-modal slides. We applied our method to ultra thin serial sections (2 μm) of a human lung tumor. In total 170 slides, stained alternately with four different stains, have been registered. Thorough visual inspection of virtual cuts through the reconstructed block perpendicular to the cutting plane shows accurate alignment of vessels and other tissue structures. This observation is confirmed by a quantitative analysis. Using nonlinear image registration, our method is able to correct locally varying deformations in tissue structures and exceeds the limitations of globally linear transformations.

Multifactorial Optimization of Contrast-Enhanced Nanofocus Computed Tomography for Quantitative Analysis of Neo-Tissue Formation in Tissue Engineering Constructs.

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Maarten Sonnaert

Full Text Available To progress the fields of tissue engineering (TE and regenerative medicine, development of quantitative methods for non-invasive three dimensional characterization of engineered constructs (i.e. cells/tissue combined with scaffolds becomes essential. In this study, we have defined the most optimal staining conditions for contrast-enhanced nanofocus computed tomography for three dimensional visualization and quantitative analysis of in vitro engineered neo-tissue (i.e. extracellular matrix containing cells in perfusion bioreactor-developed Ti6Al4V constructs. A fractional factorial 'design of experiments' approach was used to elucidate the influence of the staining time and concentration of two contrast agents (Hexabrix and phosphotungstic acid and the neo-tissue volume on the image contrast and dataset quality. Additionally, the neo-tissue shrinkage that was induced by phosphotungstic acid staining was quantified to determine the operating window within which this contrast agent can be accurately applied. For Hexabrix the staining concentration was the main parameter influencing image contrast and dataset quality. Using phosphotungstic acid the staining concentration had a significant influence on the image contrast while both staining concentration and neo-tissue volume had an influence on the dataset quality. The use of high concentrations of phosphotungstic acid did however introduce significant shrinkage of the neo-tissue indicating that, despite sub-optimal image contrast, low concentrations of this staining agent should be used to enable quantitative analysis. To conclude, design of experiments allowed us to define the most optimal staining conditions for contrast-enhanced nanofocus computed tomography to be used as a routine screening tool of neo-tissue formation in Ti6Al4V constructs, transforming it into a robust three dimensional quality control methodology.

Catecholaminergic development of fetal rat ventral mesencephalon : Characterization by high-performance liquid chromatography with electrochemical detection and immunohistochemistry

NARCIS (Netherlands)

Tomasini, R; Kema, IP; Muskiet, FAJ; Meiborg, G; Staal, MJ; Go, KG

We determined dopamine (DA), noradrenaline (NA), and adrenaline (A), as well as immunohistochemically stained tyrosine hydroxylase (TH) and DA in dissected rat ventral mesencephalon (VM) tissue from Embryonic Day (ED) 14 to Postnatal Day (P) 17. Whole VM tissue DA, NA, and A contents increased with

Endothelial cell markers in vascular neoplasms: an immunohistochemical study comparing factor VIII-related antigen, blood group specific antigens, 6-keto-PGF1 alpha, and Ulex europaeus 1 lectin.

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Little, D; Said, J W; Siegel, R J; Fealy, M; Fishbein, M C

1986-06-01

Markers for endothelial cells including Ulex europaeus 1 lectin, blood group A, B, and H, and the prostaglandin metabolite 6-keto-PGF1 alpha were evaluated in paraffin secretions from formalin-fixed benign and malignant vascular neoplasms using a variety of immunohistochemical techniques, and results compared with staining for factor VIII-related antigen. Staining for Ulex appeared more sensitive than factor VIII-related antigen in identifying poorly differentiated neoplasms including haemangiosarcomas and spindle cell proliferations in Kaposi's sarcoma. Staining for blood group related antigens correlated with blood group in all cases. Ulex europaeus 1 lectin was the only marker for endothelial cells in lymphangiomas.

Protein 53 expression in a mixed Labrador subcutaneous lymphoma

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Annahita Rezaie

2012-06-01

Full Text Available An 11 year – old mixed female Labrador was presented with two masses in trunk and neck. The tumoral masses were excised and sent for histopathological and immunohistochemical analyses. Histopathological examination of masses revealed diffuse infiltration of small sized lymphoid cells in subcutaneous tissue which were intense around the blood vessels. More than 10% lymphoid cells were CD3 positive in the immunohistochemical staining and most of them were accumulated around vessels. Protein 53 (p53 expression was detected by brown nuclei in immunohistochemical staining. Subcutaneous lymphoma was diagnosed according to histopathological results. After 6 months the case was referred with multicentric lymphoma and based on the owner request euthanasia was performed. These findings emphasize on poor prognosis for tumors with p53 mutation.

Blockface histology with optical coherence tomography: a comparison with Nissl staining.

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Magnain, Caroline; Augustinack, Jean C; Reuter, Martin; Wachinger, Christian; Frosch, Matthew P; Ragan, Timothy; Akkin, Taner; Wedeen, Van J; Boas, David A; Fischl, Bruce

2014-01-01

Spectral domain optical coherence tomography (SD-OCT) is a high resolution imaging technique that generates excellent contrast based on intrinsic optical properties of the tissue, such as neurons and fibers. The SD-OCT data acquisition is performed directly on the tissue block, diminishing the need for cutting, mounting and staining. We utilized SD-OCT to visualize the laminar structure of the isocortex and compared cortical cytoarchitecture with the gold standard Nissl staining, both qualitatively and quantitatively. In histological processing, distortions routinely affect registration to the blockface image and prevent accurate 3D reconstruction of regions of tissue. We compared blockface registration to SD-OCT and Nissl, respectively, and found that SD-OCT-blockface registration was significantly more accurate than Nissl-blockface registration. Two independent observers manually labeled cortical laminae (e.g. III, IV and V) in SD-OCT images and Nissl stained sections. Our results show that OCT images exhibit sufficient contrast in the cortex to reliably differentiate the cortical layers. Furthermore, the modalities were compared with regard to cortical laminar organization and showed good agreement. Taken together, these SD-OCT results suggest that SD-OCT contains information comparable to standard histological stains such as Nissl in terms of distinguishing cortical layers and architectonic areas. Given these data, we propose that SD-OCT can be used to reliably generate 3D reconstructions of multiple cubic centimeters of cortex that can be used to accurately and semi-automatically perform standard histological analyses. © 2013.

The Histological and Immunohistochemical Features of the Skin Lesions in CANDLE Syndrome

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Torrelo, Antonio; Colmenero, Isabel; Requena, Luis; Paller, Amy S.; Ramot, Yuval; Lee, Chyi-Chia Richard; Vera, Angel; Zlotogorski, Abraham; Goldbach-Mansky, Raphaela; Kutzner, Heinz

2015-01-01

Chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE) syndrome is a newly characterized autoinflammatory disorder, caused by mutations in PSMB8. It is characterized by early-onset fevers, accompanied by a widespread, violaceous and often annular, cutaneous eruption. While the exact pathogenesis of this syndrome is still obscure, it is postulated that the inflammatory disease manifestations stem from excess secretion of interferons. Based on preliminary blood cytokine and gene expression studies, the signature seems to come mostly from type I interferons, which are proposed to lead to the recruitment of immature myeloid cells into the dermis and subcutis. In this study, we systematically analyzed skin biopsies from 6 CANDLE syndrome patients by routine histopathology and immunohistochemistry methods. Skin lesions showed the presence of extensive mixed dermal and subcutaneous inflammatory infiltrate, composed of mononuclear cells, atypical myeloid cells, neutrophils, eosinophils and some mature lymphocytes. Positive LEDER and myeloperoxidase staining supported the presence of myeloid cells. Positive CD68/PMG1 and CD163 staining confirmed the existence of histiocytes and monocytic macrophages in the inflammatory infiltrate. CD123 staining was positive, demonstrating the presence of plasmacytoid dendritic cells. Uncovering the unique histopathologic and immunohistochemical features of CANDLE syndrome provides tools for rapid and specific diagnosis of this disorder as well as further insight into the pathogenesis of this severe, life-threatening condition. PMID:26091509

Ovarian Brenner tumors and Walthard nests: a histologic and immunohistochemical study.

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Roma, Andres A; Masand, Ramya P

2014-12-01

Brenner tumors are composed of urothelial/transitional-type epithelium and, hence, are morphologically similar to Walthard nests and tubal/mesothelial transitional metaplasia. In this study, we analyzed immunohistochemical markers on Brenner tumors to explore Müllerian as well as Wolffian and germ cell derivation. We also attempted to explore their possible association with tubal/paratubal Walthard nests/transitional metaplasia, using the same immunostains. Thirty-two consecutive cases of Brenner tumors were identified. Thirteen (43%) of the patients had Walthard nests in the tubal/periovarian soft tissue. All Brenner tumors were diffusely positive for GATA3 (strongly positive in 30/32 and weakly positive in the remaining 2) and negative for PAX8, PAX2, and SALL4. Similarly, all Walthard nests were positive for GATA3, whereas only 3 (23%) of 13 showed occasional PAX8 expression; all were negative for PAX2 and SALL4. In our study, more than 40% of Brenner tumors had associated Walthard nests. The similar morphology and immunoprofile of Brenner tumors and Walthard nests suggest a probable link between Brenner tumors and Walthard nests. Two additional cases presented highlight small transitional lesions involving the ovary: a possible precursor lesion or the initial steps of Brenner tumor formation. Brenner tumors and most Walthard nests lacked staining for Müllerian (PAX8 and PAX2) and germ cell tumor markers (SALL4). Copyright © 2014 Elsevier Inc. All rights reserved.

Transjugular Intrahepatic Portosystemic Shunt: Histologic and Immunohistochemical Study of Autopsy Cases

International Nuclear Information System (INIS)

Terayama, Noboru; Matsui, Osamu; Kadoya, Masumi; Yoshikawa, Jun; Gabata, Toshifumi; Miyayama, Shiro; Takashima, Tsutomu; Kobayashi, Kenichi; Nakanishi, Isao; Nakanuma, Yasuni

1997-01-01

Purpose: To assess the histologic findings associated with stenosed and occluded transjugular intrahepatic portosystemic shunt (TIPS) tracts. Methods: Four TIPS tracts within three autopsy livers were histologically studied for vascular components by routine staining and immunohistochemical staining. TIPS had been performed for bleeding from esophageal varices in patients with cirrhosis of the liver. Results: Two TIPS, examined on days 4 and 53, showed occlusion by fibrin thrombus. In the former, no endothelial cells were detected, but coagulative necrosis of hepatocytes was found in the surrounding liver. In the latter, bile pigments were seen on the luminal surface. In the two other TIPS without tract occlusion, examined on days 49 and 293, a layer of endothelial cells, proliferation of smooth muscle cells, and deposition of an extracellular matrix such as collagen were confirmed. In the tract examined on day 293, there was protrusion of hepatocytes into the lumen through the stent wires. Conclusion: Short- and midterm TIPS occlusions were caused by thrombus forming after necrosis of hepatocytes and bile leakage, respectively. Long-term TIPS stenosis was associated with a combination of pseudointimal hyperplasia and ingrowth of hepatocytes

leaves extracts as counter stain in gram staining reaction 56

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DR. AMINU

is a stain with color contrasting to the principal stain, making the stained ... technology today, the Gram's staining method remains ... was aimed at employing the use of Henna leaves extract as ... fragrant, white or rose flowers in clusters. It is.

Development of a monoclonal antibody that specifically detects tissue inhibitor of metalloproteinase-4 (TIMP-4) in formalin-fixed, paraffin-embedded human tissues.

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Donover, P Scott; Wojciechowski, Brian S; Thirumaran, Rajesh; Zemba-Palko, Vlasta; Prendergast, George C; Wallon, U Margaretha

2010-08-01

Overexpression of the extracellular metalloproteinase inhibitor TIMP-4 in estrogen receptor-negative breast cancers was found recently to be associated with a poor prognosis for survival. To pursue exploration of the theranostic applications of TIMP-4, specific antibodies with favorable properties for immunohistochemical use and other clinical assays are needed. Here we report the characterization of a monoclonal antibody (clone 9:4-7) specific for full-length human TIMP-4 with suitable qualities. The antibody was determined to be an IgG(2b) immunoglobulin. In enzyme-linked immunosorbent assay (ELISA) and immunoblotting assays, it did not exhibit any detectable crossreactivity with recombinant forms of the other human TIMPs 1, 2, and 3. In contrast, the antibody displayed high specificity and sensitivity for TIMP-4 including in formalin-fixed and paraffin-embedded specimens of human breast specimens. An analysis of tissue microarrays of human cancer and corresponding normal tissues revealed specific staining patterns with excellent signal-to-noise ratios. This study documents TIMP-4 monoclonal antibody clone 9:4-7 as an effective tool for preclinical and clinical investigations. Published 2010 Wiley-Liss, Inc.

Digital immunohistochemistry platform for the staining variation monitoring based on integration of image and statistical analyses with laboratory information system.

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Laurinaviciene, Aida; Plancoulaine, Benoit; Baltrusaityte, Indra; Meskauskas, Raimundas; Besusparis, Justinas; Lesciute-Krilaviciene, Daiva; Raudeliunas, Darius; Iqbal, Yasir; Herlin, Paulette; Laurinavicius, Arvydas

2014-01-01

Digital immunohistochemistry (IHC) is one of the most promising applications brought by new generation image analysis (IA). While conventional IHC staining quality is monitored by semi-quantitative visual evaluation of tissue controls, IA may require more sensitive measurement. We designed an automated system to digitally monitor IHC multi-tissue controls, based on SQL-level integration of laboratory information system with image and statistical analysis tools. Consecutive sections of TMA containing 10 cores of breast cancer tissue were used as tissue controls in routine Ki67 IHC testing. Ventana slide label barcode ID was sent to the LIS to register the serial section sequence. The slides were stained and scanned (Aperio ScanScope XT), IA was performed by the Aperio/Leica Colocalization and Genie Classifier/Nuclear algorithms. SQL-based integration ensured automated statistical analysis of the IA data by the SAS Enterprise Guide project. Factor analysis and plot visualizations were performed to explore slide-to-slide variation of the Ki67 IHC staining results in the control tissue. Slide-to-slide intra-core IHC staining analysis revealed rather significant variation of the variables reflecting the sample size, while Brown and Blue Intensity were relatively stable. To further investigate this variation, the IA results from the 10 cores were aggregated to minimize tissue-related variance. Factor analysis revealed association between the variables reflecting the sample size detected by IA and Blue Intensity. Since the main feature to be extracted from the tissue controls was staining intensity, we further explored the variation of the intensity variables in the individual cores. MeanBrownBlue Intensity ((Brown+Blue)/2) and DiffBrownBlue Intensity (Brown-Blue) were introduced to better contrast the absolute intensity and the colour balance variation in each core; relevant factor scores were extracted. Finally, tissue-related factors of IHC staining variance were

Immunohistochemical Analysis of Scarring Trachoma Indicates Infiltration by Natural Killer and Undefined CD45 Negative Cells.

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Hu, Victor H; Luthert, Philip J; Derrick, Tamsyn; Pullin, James; Weiss, Helen A; Massae, Patrick; Mtuy, Tara; Makupa, William; Essex, David; Mabey, David C W; Bailey, Robin L; Holland, Martin J; Burton, Matthew J

2016-05-01

The phenotype and function of immune cells infiltrating the conjunctiva in scarring trachoma have yet to be fully characterized. We assessed tissue morphology and immunophenotype of cellular infiltrates found in trachomatous scarring compared to control participants. Clinical assessments and conjunctival biopsy samples were obtained from 34 individuals with trachomatous scarring undergoing trichiasis surgery and 33 control subjects undergoing cataract or retinal detachment surgery. Biopsy samples were fixed in buffered formalin and embedded in paraffin wax. Hematoxylin and eosin (H&E) staining was performed for assessment of the inflammatory cell infiltrate. Immunohistochemical staining of single markers on individual sections was performed to identify cells expressing CD3 (T-cells), CD4 (helper T-cells), CD8 (suppressor/cytotoxic T-cells and Natural Killer, NK, cells), NCR1 (NK cells), CD20 (B-cells), CD45 (nucleated hematopoietic cells), CD56 (NK and T-cells), CD68 (macrophages/monocytes) and CD83 (mature dendritic cells). The degree of scarring was assessed histologically using cross-polarized light to visualize collagen fibres. Scarring, regardless of clinical inflammation, was associated with increased inflammatory cell infiltrates on H&E and CD45 staining. Scarring was also associated with increased CD8+ and CD56+ cells, but not CD3+ cells, suggestive of a NK cell infiltrate. This was supported by the presence of NCR1+ cells. There was some increase in CD20+ cells, but no evidence for increased CD4+, CD68+ or CD83+ cells. Numerous CD45 negative cells were also seen in the population of infiltrating inflammatory cells in scarred conjunctiva. Disorganization of the normal collagen architecture was strongly associated with clinical scarring. These data point to the infiltration of immune cells with a phenotype suggestive of NK cells in conjunctival trachomatous scarring. A large proportion of CD45 negative inflammatory cells were also present. Future work should

Reevaluation and reclassification of resected lung carcinomas originally diagnosed as squamous cell carcinoma using immunohistochemical analysis

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Kadota, Kyuichi; Nitadori, Jun-ichi; Rekhtman, Natasha; Jones, David R.; Adusumilli, Prasad S.; Travis, William D.

2015-01-01

Currently, non-small cell lung carcinomas are primarily classified by light microscopy. However, recent studies have shown that poorly-differentiated tumors are more accurately classified by immunohistochemistry. In this study, we investigated the use of immunohistochemical analysis in reclassifying lung carcinomas that were originally diagnosed as squamous cell carcinoma. Tumor slides and blocks were available for histologic evaluation, and tissue microarrays were constructed from 480 patients with resected lung carcinomas originally diagnosed as squamous cell carcinoma between 1999 and 2009. Immunohistochemistry for p40, p63, thyroid transcription factor-1 (TTF-1; clone SPT24 and 8G7G3/1), Napsin A, Chromogranin A, Synaptophysin, and CD56 were performed. Staining intensity (weak, moderate, or strong) and distribution (focal or diffuse) were also recorded. Of all, 449 (93.5%) patients were confirmed as having squamous cell carcinomas; the cases were mostly diffusely positive for p40 and negative for TTF-1 (8G7G3/1). Twenty cases (4.2%) were reclassified as adenocarcinoma since they were positive for TTF-1 (8G7G3/1 or SPT24) with either no or focal p40 expression, and all of them were poorly-differentiated with squamoid morphology. In addition, 1 case was reclassified as adenosquamous carcinoma, 4 cases as large cell carcinoma, 4 cases as large cell neuroendocrine carcinoma, and 2 cases as small cell carcinoma. In poorly-differentiated non-small cell lung carcinomas, an accurate distinction between squamous cell carcinoma and adenocarcinoma cannot be reliably determined by morphology alone and requires immunohistochemical analysis, even in resected specimens. Our findings suggest that TTF-1 8G7G3/1 may be better suited as the primary antibody in differentiating adenocarcinoma from squamous cell carcinoma. PMID:25871623

Ectopic Hard Tissue Formation by Odonto/Osteogenically In Vitro Differentiated Human Deciduous Teeth Pulp Stem Cells.

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Kim, Seunghye; Song, Je Seon; Jeon, Mijeong; Shin, Dong Min; Kim, Seong-Oh; Lee, Jae Ho

2015-07-01

There have been many attempts to use the pulp tissue from human deciduous teeth for dentin or bone regeneration. The objective of this study was to determine the effects of odonto/osteogenic in vitro differentiation of deciduous teeth pulp stem cells (DTSCs) on their in vivo hard tissue-forming potential. DTSCs were isolated from extracted deciduous teeth using the outgrowth method. These cells were exposed to odonto/osteogenic stimuli for 4 and 8 days (Day 4 and Day 8 groups, respectively), while cells in the control group were cultured in normal medium. The in vitro differentiated DTSCs and the control DTSCs were transplanted subcutaneously into immunocompromised mice with macroporous biphasic calcium phosphate and sacrificed at 8 weeks post-implantation. The effect of odonto/osteogenic in vitro differentiation was evaluated using alkaline phosphatase (ALP) staining and quantitative reverse transcription polymerase chain reaction (RT-PCR). The in vivo effect was evaluated by qualitative RT-PCR, assessment of ALP activity, histologic analysis, and immunohistochemical staining. The amount of hard tissue was greater in Day 4 group than Day 8 group (p = 0.014). However, Day 8 group generated lamellar bone-like structure, which was immunonegative to anti-human dentin sialoprotein with significantly low expression level of DSPP compared with the control group (p = 0.008). This study demonstrates that odonto/osteogenic in vitro differentiation of DTSCs enhances the formation of bone-like tissue, instead of dentin-like tissue, when transplanted subcutaneously using MBCP as a carrier. The odonto/osteogenic in vitro differentiation of DTSCs may be an effective modification that enhances in vivo bone formation by DTSCs.

Comparative immunohistochemical expression of β-catenin, EGFR, ErbB2, and p63 in adamantinomatous and papillary craniopharyngiomas

International Nuclear Information System (INIS)

Eshebaa, Gh.E.; Hassan, A.A.

2015-01-01

Craniopharyngiomas (CPs) are rare epithelial tumors located mainly in the sellar/parasellar region. CPs have been classified into histopathologically, genetically, clinically and prognostically two distinctive subtypes: adamantinomatous and papillary variants. Aim To determine the immunohistochemical expression of β-catenin, EGFR, ErbB2, and p63 in adamantinomatous and papillary CPs. Materials and methods β-Catenin, EGFR, ErbB2, and p63 immunostaining was performed on paraffin embedded tissue sections of 25 CPs including 18 adamantinomatous craniopharyngioma (ACP) and 7 cases of papillary craniopharyngiomas (PCPs). Results 17 cases (94%) of ACP exhibited strong nuclear/cytoplasmic expression of β-catenin. On the contrary, all cases of PCP showed exclusively membranous expression (ρ value <0.0001). Regarding EGFR, 15 (83%) and 5 cases (71%) of APC and PCP respectively were positive. On the other hand, only 3 cases (17%) of APC and none of PCP exhibited positivity for ErbB2. p63 over-expression was observed in 16 cases of ACP (89%) and 6 cases of PCP (86%). However, the distribution of p63 staining was diffuse in ACP, while in PCP; the staining was mainly restricted to the basal cell layer. Conclusion Nuclear accumulation of β-catenin is a diagnostic hallmark of the ACP and is very helpful in the differential diagnosis between both ACP and PCP in the setting of small biopsies. Moreover, the restricted nuclear β-catenin accumulation in the cohesive cell clusters within the whorl-like areas supports that aberrant β-catenin expression may play a role in the morphogenesis of ACP.

Twist and YB-1 gene expression in cervical cancer and precancerous tissue and their correlation with cell invasion

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Qin Tian

2017-04-01

Full Text Available Objective: To study the correlation of Twist and YB-1 gene expression in cervical cancer and precancerous tissue with cell invasion. Methods: Cervical cancer tissue, precancerous tissue and normal cervical tissue surgically removed in our hospital between May 2013 and April 2015 were collected; immunohistochemical staining kits were used to detect the positive protein expression rate of Twist and YB-1 gene; fluorescence quantitative PCR kits were used to detect Twist, YB-1 and invasion gene mRNA expression. Results: Twist and YB-1 mRNA expression and positive protein expression rate as well as USP22, Rab11, Rac1 and ANXA5 mRNA expression in cervical cancer tissue and precancerous tissue were significantly higher than those in normal cervical tissue, Twist and YB-1 mRNA expression and positive protein expression rate as well as USP22, Rab11, Rac1 and ANXA5 mRNA expression in cervical cancer tissue were significantly higher than those in precancerous tissue; USP22, Rab11, Rac1 and ANXA5 mRNA expression in cervical cancer tissue and precancerous tissue with positive Twist and YB-1 expression were significantly higher than those in cervical cancer tissue and precancerous tissue with negative Twist and YB-1 expression. Conclusion: Highly expressed Twist and YB-1 in cervical cancer and precancerous tissue can promote cell invasion.

Basal cell carcinoma vs basaloid squamous cell carcinoma of the skin: an immunohistochemical reappraisal.

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Webb, David V; Mentrikoski, Mark J; Verduin, Lindsey; Brill, Louis B; Wick, Mark R

2015-04-01

Typical cutaneous basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) are morphologically dissimilar. It is well known, however, that poorly differentiated SCC may assume a basaloid phenotype, complicating the histologic distinction between these 2 neoplasms. Selected immunohistochemical stains have been used in the past to aid in that differential diagnosis. In the current study, additional markers were evaluated to determine whether they would be helpful in that regard. Twenty-nine cases of metatypical (squamoid) BCC (MBCC) and 25 examples of basaloid SCC (BSCC) were studied using the antibodies Ber-EP4 and MOC-31 as well as a plant lectin preparation from Ulex europaeus I (UEA-1). The resulting immunostains were interpreted independently by 3 pathologists, and the results showed that MBCCs demonstrated strong and diffuse staining for Ber-EP4 (25/29) and MOC-31 (29/29). In contrast, BSCCs tended to be only sporadically reactive for both markers (4/25 and 1/25 cases, respectively). Labeling for UEA-1 was observed in almost all BSCCs (24/25), but only 6 of 29 cases of MBCC showed limited, focal staining with that lectin. These data suggest that MOC-31 is a useful marker in the specified differential diagnosis, especially when used together with UEA-1. Copyright © 2015 Elsevier Inc. All rights reserved.

Port-Wine Stains

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... Safe Videos for Educators Search English Español Port-Wine Stains KidsHealth / For Parents / Port-Wine Stains What's ... Manchas de vino de oporto What Are Port-Wine Stains? A port-wine stain is a type ...

Experimental study of tissue-engineered cartilage allograft with RNAi chondrocytes in vivo

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Wang ZH

2014-05-01

Full Text Available Zhenghui Wang,1 Xiaoli Li,2 Xi-Jing He,3 Xianghong Zhang,1 Zhuangqun Yang,4 Min Xu,1 Baojun Wu,1 Junbo Tu,5 Huanan Luo,1 Jing Yan11Department of Otolaryngology – Head and Neck Surgery, 2Department of Dermatology, 3Department of Orthopedics, The Second Hospital, Xi’an Jiaotong University, 4Department of Plastic and Burns Surgery, The First Hospital, Xi’an Jiaotong University, 5Department of Oral and Maxillofacial Plastic Surgery, The Stomatological Hospital, Xi’an Jiaotong University, Xi’an, People’s Republic of ChinaPurpose: To determine the effects of RNA interference (RNAi on chondrocyte proliferation, function, and immunological rejection after allogenic tissue-engineered cartilage transplantation within bone matrix gelatin scaffolds.Methods: Seven million rat normal and RNAi chondrocytes were harvested and separately composited with fibrin glue to make the cell suspension, and then transplanted subcutaneously into the back of Sprague Dawley rats after being cultured for 10 days in vitro. Untransplanted animals served as the control group. The allograft and immunological response were examined at 1, 2, 4, 8, and 12 months postoperatively with hematoxylin and eosin histochemical staining, immunohistochemical staining (aggrecan, type II collagen, class I and II major histocompatibility complex, and flow cytometry for peripheral blood cluster of differentiation 4+ (CD4+ and CD8+ T-cells.Results: There was no infection or death in the rats except one, which died in the first week. Compared to the control group, the RNAi group had fewer eukomonocytes infiltrated, which were only distributed around the graft. The ratio of CD4+/CD8+ T-cells in the RNAi group was significantly lower than the normal one (P<0.05. There were many more positively stained chondrocytes and positively stained areas around the cells in the RNAi group, which were not found in the control group.Conclusion: The aggrecanase-1 and aggrecanase-2 RNAi for chondrocytes

A prototype for unsupervised analysis of tissue microarrays for cancer research and diagnostics.

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Chen, Wenjin; Reiss, Michael; Foran, David J

2004-06-01

The tissue microarray (TMA) technique enables researchers to extract small cylinders of tissue from histological sections and arrange them in a matrix configuration on a recipient paraffin block such that hundreds can be analyzed simultaneously. TMA offers several advantages over traditional specimen preparation by maximizing limited tissue resources and providing a highly efficient means for visualizing molecular targets. By enabling researchers to reliably determine the protein expression profile for specific types of cancer, it may be possible to elucidate the mechanism by which healthy tissues are transformed into malignancies. Currently, the primary methods used to evaluate arrays involve the interactive review of TMA samples while they are viewed under a microscope, subjectively evaluated, and scored by a technician. This process is extremely slow, tedious, and prone to error. In order to facilitate large-scale, multi-institutional studies, a more automated and reliable means for analyzing TMAs is needed. We report here a web-based prototype which features automated imaging, registration, and distributed archiving of TMAs in multiuser network environments. The system utilizes a principal color decomposition approach to identify and characterize the predominant staining signatures of specimens in color space. This strategy was shown to be reliable for detecting and quantifying the immunohistochemical expression levels for TMAs.

Delineation of Chondroid Lipoma: An Immunohistochemical and Molecular Biological Analysis

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de Vreeze, Ronald S. A.; van Coevorden, Frits; Boerrigter, Lucie; Nederlof, Petra M.; Haas, Rick L.; Bras, Johannes; Rosenwald, Andreas; Mentzel, Thomas; de Jong, Daphne

2011-01-01

Aims. Chondroid lipoma (CL) is a benign tumor that mimics a variety of soft tissue tumors and is characterized by translocation t(11;16). Here, we analyze CL and its histological mimics. Methods. CL (n = 4) was compared to a variety of histological mimics (n = 83) for morphological aspects and immunohistochemical features including cyclinD1(CCND1). Using FISH analysis, CCND1 and FUS were investigated as potential translocation partners. Results. All CLs were strongly positive for CCND1. One of 4 myoepitheliomas, CCND1, was positive. In well-differentiated lipomatous tumors and in chondrosarcomas, CCND1 was frequently expressed, but all myxoid liposarcomas were negative. FISH analysis did not give support for direct involvement of CCND1 and FUS as translocation partners. Conclusions. Chondroid lipoma is extremely rare and has several and more prevalent histological mimics. The differential diagnosis of chondroid lipomas can be unraveled using immunohistochemical and molecular support. PMID:21559269

Expression of E-cadherin and involucrin in leukoplakia and oral cancer: an immunocytochemical and immunohistochemical study

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Alessandra Dutra da SILVA

2017-03-01

Full Text Available Abstract To assess the immunocytochemical and immunohistochemical correlation of adhesion (E-cadherin and cell differentiation (involucrin molecules in oral leukoplakia and oral squamous cell carcinoma. Cytological samples and biopsies were obtained from male and female patients aged over 30 years with oral leukoplakia (n = 30 and oral squamous cell carcinoma (n = 22. Cell scrapings and the biopsy were performed at the site of the lesion and histological slides were prepared for the immunocytochemical analysis of exfoliated oral mucosal cells and for the immunohistochemical analysis of biopsy tissues using E-cadherin and involucrin. Spearman’s correlation and kappa coefficients were used to assess the correlation and level of agreement between the techniques. Immunostaining for E-cadherin and involucrin by both techniques was similar in the superficial layers of the histological sections compared with cell scrapings. However, there was no statistical correlation and agreement regarding the immunocytochemical and immunohistochemical expression of E-cadherin and involucrin in oral leukoplakia (R = 0.01, p = 0.958 (Kappa = 0.017, p = 0.92 or in oral squamous cell carcinoma (R = 0.26, p = 0.206 (Kappa = 0.36, p = 0.07. The immunoexpression of E-cadherin and involucrin in tissues is consistent with the expression patterns observed in exfoliated oral mucosal cells, despite the lack of a statistically significant correlation. There is an association of the histopathological characteristics of leukoplakia with the expression E-cadherin and of the microscopic aspects of oral squamous cell carcinoma with immunohistochemical expression of involucrin.

Study of polarization colors in the connective tissue wall of odontogenic cysts using picrosirius red stain

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Anusha Shetty

2015-01-01

Full Text Available Background: Lesions of odontogenic origin comprise the heterogeneous group ranging from hamartomatous proliferations, cysts to benign and malignant tumors. Interplay between the epithelium and connective tissue can be assumed to play a significant role in the pathogenesis of odontogenic cysts. Aims and Objectives: A study was taken up to show the role of picrosirius red (PSR stain to demonstrate the fibers and also to assess the difference in the nature of the fibers (different color patterns and to find out the role of it, if any in the pathogenesis and biological behavior of the commonly occurring odontogenic cysts. Materials and Methods: Collagen fibers of 30 cases of odontogenic cysts (10 radicular cysts, 10 odontogenic keratocysts (OKC′s, and 10 dentigerous cysts were studied by staining the sections with PSR stain and examining them under bright field and polarizing microscope. Results: Sixty-seven percentage of the thin collagen fibers and 55% of the thick fibers in radicular cyst showed green-yellow birefringence. Fifty-seven percentage of the thin collagen fibers and 15% of the thick fibers in OKC showed green-yellow birefringence. Eighty-two percentage of the thin collagen fibers and 66% of the thick fibers in dentigerous cysts showed green-yellow birefringence. Rest of the fibers showed orange-red birefringence. Statistical analysis with one-way ANOVA was significant with a P < 0.01 only for thick fibers. Moreover, comparison of polarization colors of thick fibers of odontogenic cysts with duration of the lesion gave statistically significant results. Conclusion: The observations in the present study with respect to color profiles of the collagen fibers in the three commonly occurring odontogenic cysts possibly explain the biological behavior of the lesions. The predominant orange-red birefringence in OKC′s in comparison to radicular and dentigerous cysts suggests that OKC′s exhibit well organized and tightly packed fibers. This

Sarcoglycan complex in masseter and sternocleidomastoid muscles of baboons: an immunohistochemical study

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G. Cutroneo

2015-06-01

Full Text Available The sarcoglycan complex consists of a group of single-pass transmembrane glycoproteins that are essential to maintain the integrity of muscle membranes. Any mutation in each sarcoglycan gene causes a series of recessive autosomal dystrophin-positive muscular dystrophies. Negative fibres for sarcoglycans have never been found in healthy humans and animals. In this study, we have investigated whether the social ranking has an influence on the expression of sarcoglycans in the skeletal muscles of healthy baboons. Biopsies of masseter and sternocleidomastoid muscles were processed for confocal immunohistochemical detection of sarcoglycans. Our findings showed that baboons from different social rankings exhibited different sarcoglycan expression profiles. While in dominant baboons almost all muscles were stained for sarcoglycans, only 55% of muscle fibres showed a significant staining. This different expression pattern is likely to be due to the living conditions of these primates. Sarcoglycans which play a key role in muscle activity by controlling contractile forces may influence the phenotype of muscle fibres, thus determining an adaptation to functional conditions. We hypothesize that this intraspecies variation reflects an epigenetic modification of the muscular protein network that allows baboons to adapt progressively to a different social status.

A Historical Perspective on the Identification of Cell Types in Pancreatic Islets of Langerhans by Staining and Histochemical Techniques.

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Baskin, Denis G

2015-08-01

Before the middle of the previous century, cell types of the pancreatic islets of Langerhans were identified primarily on the basis of their color reactions with histological dyes. At that time, the chemical basis for the staining properties of islet cells in relation to the identity, chemistry and structure of their hormones was not fully understood. Nevertheless, the definitive islet cell types that secrete glucagon, insulin, and somatostatin (A, B, and D cells, respectively) could reliably be differentiated from each other with staining protocols that involved variations of one or more tinctorial techniques, such as the Mallory-Heidenhain azan trichrome, chromium hematoxylin and phloxine, aldehyde fuchsin, and silver impregnation methods, which were popularly used until supplanted by immunohistochemical techniques. Before antibody-based staining methods, the most bona fide histochemical techniques for the identification of islet B cells were based on the detection of sulfhydryl and disulfide groups of insulin. The application of the classical islet tinctorial staining methods for pathophysiological studies and physiological experiments was fundamental to our understanding of islet architecture and the physiological roles of A and B cells in glucose regulation and diabetes. © The Author(s) 2015.

Retrospective study of 338 canine oral melanomas with clinical, histologic, and immunohistochemical review of 129 cases.

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Ramos-Vara, J A; Beissenherz, M E; Miller, M A; Johnson, G C; Pace, L W; Fard, A; Kottler, S J

2000-11-01

Diagnostic records from 338 canine oral melanomas in 338 dogs received at the Veterinary Medical Diagnostic Laboratory (1992-1999) were reviewed. Of these tumors, 122 plus an additional 7 metastatic melanomas of unknown origin were selected for clinical follow-up, histologic review, and immunohistochemistry. Chow Chow, Golden Retriever, and Pekingese/Poodle mix breeds were overrepresented, whereas Boxer and German Shepherd breeds were underrepresented. There was no gender predisposition and the average age at presentation was 11.4 years. Forty-nine dogs were euthanized due to recurrence or metastasis. The average postsurgical survival time was 173 days. The gingiva and the labial mucosa were the most common sites. Most tumors were composed of either polygonal cells (27 cases, 20.9%), spindle cells (44 cases, 34.1%), or a mixture of the two (polygonal and spindle) (54 cases, 41.9%). Clear cell (3 cases, 2.3%) and adenoid/papillary (1 case, 0.8%) patterns were uncommon. The metastases of 6/6 oral melanomas had morphologic and immunohistochemical features similar to those of the primary tumors. Immunohistochemically, Melan A was detected in 113/122 oral (92.6%) and 5/7 (71.9%) metastatic melanomas. Only 4/163 nonmelanocytic tumors were focally and weakly positive for Melan A. Antibodies against vimentin, S100 protein, and neuron-specific enolase stained 129 (100%), 98 (76%), and 115 (89.1%) of 129 melanomas, respectively. Antibodies against other melanocytic-associated antigens (tyrosinase, glycoprotein 100) did not yield adequate staining. We conclude that Melan A is a specific and sensitive marker for canine melanomas.

Immunohistochemical assessment of NY-ESO-1 expression in esophageal adenocarcinoma resection specimens.

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Hayes, Stephen J; Hng, Keng Ngee; Clark, Peter; Thistlethwaite, Fiona; Hawkins, Robert E; Ang, Yeng

2014-04-14

To assess NY-ESO-1 expression in a cohort of esophageal adenocarcinomas. A retrospective search of our tissue archive for esophageal resection specimens containing esophageal adenocarcinoma was performed, for cases which had previously been reported for diagnostic purposes, using the systematised nomenclature of human and veterinary medicine coding system. Original haematoxylin and eosin stained sections were reviewed, using light microscopy, to confirm classification and tumour differentiation. A total of 27 adenocarcinoma resection specimens were then assessed using immunohistochemistry for NY-ESO-1 expression: 4 well differentiated, 14 moderately differentiated, 4 moderate-poorly differentiated, and 5 poorly differentiated. Four out of a total of 27 cases of esophageal adenocarcinoma examined (15%) displayed diffuse cytoplasmic and nuclear expression for NY-ESO-1. They displayed a heterogeneous and mosaic-type pattern of diffuse staining. Diffuse cytoplasmic staining was not identified in any of these structures: stroma, normal squamous epithelium, normal submucosal gland and duct, Barrett's esophagus (goblet cell), Barrett's esophagus (non-goblet cell) and high grade glandular dysplasia. All adenocarcinomas showed an unexpected dot-type pattern of staining at nuclear, paranuclear and cytoplasmic locations. Similar dot-type staining, with varying frequency and size of dots, was observed on examination of Barrett's metaplasia, esophageal submucosal gland acini and the large bowel negative control, predominantly at the crypt base. Furthermore, a prominent pattern of apical (luminal) cytoplasmic dot-type staining was observed in some cases of Barrett's metaplasia and also adenocarcinoma. A further morphological finding of interest was noted on examination of haematoxylin and eosin stained sections, as aggregates of lymphocytes were consistently noted to surround submucosal glands. We have demonstrated for the first time NY-ESO-1 expression by esophageal

Ultra-small particles of iron oxide as peroxidase for immunohistochemical detection

International Nuclear Information System (INIS)

Wu Yihang; Song Mengjie; Zhang Xiaoqing; Zhang Yu; Wang Chunyu; Gu Ning; Xin Zhuang; Li Suyi

2011-01-01

Dimercaptosuccinic acid (DMSA) modified ultra-small particles of iron oxide (USPIO) were synthesized through a two-step process. The first step: oleic acid (OA) capped Fe 3 O 4 (OA-USPIO) were synthesized by a novel oxidation coprecipitation method in H 2 O/DMSO mixing system, where DMSO acts as an oxidant simultaneously. The second step: OA was replaced by DMSA to obtain water-soluble nanoparticles. The as-synthesized nanoparticles were characterized by TEM, FTIR, TGA, VSM, DLS, EDS and UV-vis. Hydrodynamic sizes and Peroxidase-like catalytic activity of the nanoparticles were investigated. The hydrodynamic sizes of the nanoparticles (around 24.4 nm) were well suited to developing stable nanoprobes for bio-detection. The kinetic studies were performed to quantitatively evaluate the catalytic ability of the peroxidase-like nanoparticles. The calculated kinetic parameters indicated that the DMSA-USPIO possesses high catalytic activity. Based on the high activity, immunohistochemical experiments were established: using low-cost nanoparticles as the enzyme instead of expensive HRP, Nimotuzumab was conjugated onto the surface of the nanoparticles to construct a kind of ultra-small nanoprobe which was employed to detect epidermal growth factor receptor (EGFR) over-expressed on the membrane of esophageal cancer cell. The proper sizes of the probes and the result of membranous immunohistochemical staining suggest that the probes can be served as a useful diagnostic reagent for bio-detection.

Immunohistochemical study of doublecortin and nucleostemin in canine brain

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E. De Nevi

2013-03-01

Full Text Available Finding a marker of neural stem cells remains a medical research priority. It was reported that the proteins doublecortin and nucleostemin were related with stem/progenitor cells in central nervous system. The aim of the present immunohistochemical study was to evaluate the expression of these proteins and their pattern of distribution in canine brain, including age-related changes, and in non-nervous tissues. We found that doublecortin had a more specific expression pattern, related with neurogenesis and neuronal migration, while nucleostemin was expressed in most cells of almost every tissue studied. The immunolabeling of both proteins decreased with age. We may conclude that nucleostemin is not a specific marker of stem/progenitor cells in the dog. Doublecortin, however, is not an exclusive marker of neural stem cells, but also of neuronal precursors.

HISTOLOGIC, IMMUNOHISTOCHEMICAL, AND ELECTRON MICROSCOPIC CHARACTERIZATION OF A MALIGNANT IRIDOPHOROMA IN A DWARF BEARDED DRAGON (POGONA HENRYLAWSONI).

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de Brot, Simone; Sydler, Titus; Nufer, Lisbeth; Ruetten, Maja

2015-09-01

A dwarf bearded dragon (Pogona henrylawsoni) was presented with a white subcutaneous mandibular mass and multiple nodules in the oral mucosa, heart, liver, kidney, intestine, and visceral fat. Histologically, the tumor consisted of densely packed spindle-shaped cells with brow intracytoplasmic pigment that exhibited white-blue birefringence with polarized light. Immunohistochemical staining was negative for S-100 and weakly positive with melan A. Electron microscopic examination revealed cytoplasmic irregular and oblong empty spaces, laminated and often arranged into short stacks, compatible with reflecting platelet profiles typically seen in iridophores. However, in unstained ultrathin sections, electron-dense crystalline material was present, which filled the empty spaces described for stained sections before. Based on histology, immunohistochemistry, and biologic behavior, a malignant iridophoroma was diagnosed. To the authors' knowledge, iridophoromas in lizards have rarely been characterized by using electronic microscopy. Moreover, this is the first description of an iridophoroma in a dwarf bearded dragon.

Combined histochemical staining, RNA amplification, regional, and single cell cDNA analysis within the hippocampus.

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Ginsberg, Stephen D; Che, Shaoli

2004-08-01

The use of five histochemical stains (cresyl violet, thionin, hematoxylin & eosin, silver stain, and acridine orange) was evaluated in combination with an expression profiling paradigm that included regional and single cell analyses within the hippocampus of post-mortem human brains and adult mice. Adjacent serial sections of human and mouse hippocampus were labeled by histochemistry or neurofilament immunocytochemistry. These tissue sections were used as starting material for regional and single cell microdissection followed by a newly developed RNA amplification procedure (terminal continuation (TC) RNA amplification) and subsequent hybridization to custom-designed cDNA arrays. Results indicated equivalent levels of global hybridization signal intensity and relative expression levels for individual genes for hippocampi stained by cresyl violet, thionin, and hematoxylin & eosin, and neurofilament immunocytochemistry. Moreover, no significant differences existed between the Nissl stains and neurofilament immunocytochemistry for individual CA1 neurons obtained via laser capture microdissection. In contrast, a marked decrement was observed in adjacent hippocampal sections stained for silver stain and acridine orange, both at the level of the regional dissection and at the CA1 neuron population level. Observations made on the cDNA array platform were validated by real-time qPCR using primers directed against beta-actin and glyceraldehyde-3 phosphate dehydrogenase. Thus, this report demonstrated the utility of using specific Nissl stains, but not stains that bind RNA species directly, in both human and mouse brain tissues at the regional and cellular level for state-of-the-art molecular fingerprinting studies.

Hormonal receptors and vascular endothelial growth factor in juvenile nasopharyngeal angiofibroma: immunohistochemical and tissue microarray analysis.

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Liu, Zhuofu; Wang, Jingjing; Wang, Huan; Wang, Dehui; Hu, Li; Liu, Quan; Sun, Xicai

2015-01-01

This work demonstrated that juvenile nasopharyngeal angiofibromas (JNAs) express high levels of hormone receptors and vascular endothelial growth factor (VEGF) compared with normal nasal mucosa. The interaction between hormone receptors and VEGF may be involved in the initiation and growth of JNA. JNA is a rare benign tumor that occurs almost exclusively in male adolescents. Although generally regarded as a hormone-dependent tumor, this has not been proven in previous studies. The aim of this study was to investigate the role of hormone receptors in JNA and the relationship with clinical characteristics. Standard immunohistochemical microarray analysis was performed on 70 JNA samples and 10 turbinate tissue samples. Specific antibodies for androgen receptor (AR), estrogen receptor-α (ER-α), estrogen receptor-β (ER-β), progesterone receptor (PR), and VEGF were examined, and the relationships of receptor expression with age, tumor stage, and bleeding were evaluated. RESULTS showed that JNA expressed ER-α (92.9%), ER-β (91.4%), AR (65.7%), PR (12.8%), and VEGF (95.7%) at different levels. High level of VEGF was linked to elevated ER-α and ER-β. There was no significant relationship between hormonal receptors and age at diagnosis, tumor stage or bleeding. However, overexpression of ER-α was found to be an indicator of poor prognosis (p = 0.031).

Mapping stain distribution in pathology slides using whole slide imaging

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Fang-Cheng Yeh

2014-01-01

Full Text Available Background: Whole slide imaging (WSI offers a novel approach to digitize and review pathology slides, but the voluminous data generated by this technology demand new computational methods for image analysis. Materials and Methods: In this study, we report a method that recognizes stains in WSI data and uses kernel density estimator to calculate the stain density across the digitized pathology slides. The validation study was conducted using a rat model of acute cardiac allograft rejection and another rat model of heart ischemia/reperfusion injury. Immunohistochemistry (IHC was conducted to label ED1 + macrophages in the tissue sections and the stained slides were digitized by a whole slide scanner. The whole slide images were tessellated to enable parallel processing. Pixel-wise stain classification was conducted to classify the IHC stains from those of the background and the density distribution of the identified IHC stains was then calculated by the kernel density estimator. Results: The regression analysis showed a correlation coefficient of 0.8961 between the number of IHC stains counted by our stain recognition algorithm and that by the manual counting, suggesting that our stain recognition algorithm was in good agreement with the manual counting. The density distribution of the IHC stains showed a consistent pattern with those of the cellular magnetic resonance (MR images that detected macrophages labeled by ultrasmall superparamagnetic iron-oxide or micron-sized iron-oxide particles. Conclusions: Our method provides a new imaging modality to facilitate clinical diagnosis. It also provides a way to validate/correlate cellular MRI data used for tracking immune-cell infiltration in cardiac transplant rejection and cardiac ischemic injury.

The relationship of platinum resistance and ERCC1 protein expression in epithelial ovarian cancer

DEFF Research Database (Denmark)

Steffensen, Karina Dahl; Waldstrøm, Marianne; Jakobsen, Anders

2009-01-01

: Formalin-fixed, paraffin-embedded tissue sections from 101 patients with newly diagnosed ovarian cancer were used for immunohistochemical staining for the ERCC1 protein. All patients received carboplatin-paclitaxel combination chemotherapy. RESULTS: Excision repair cross-complementation group 1 enzyme...

Primary Ewing's Sarcoma of Frontoparietal Bone with Major Soft Tissue Extension: An Unusual Presentation and Review of the Literature

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Anshu Gupta

2012-01-01

Full Text Available An 11-year-old girl presented with progressively increasing swelling in scalp of 8-month duration with no neurological deficit. Local examination showed a hard swelling that seemed to be arising from frontal bone. General and systemic examination was normal. MRI revealed a well-defined lytic lesion in left frontoparietal bone with a subgaleal component. The patient was operated upon and excision of tumor with reconstruction of skull was done. Histopathological examination showed a monomorphic small round cell tumor of bone infiltrating into the subcutaneous tissue. Immunohistochemical stain showed diffuse immunopositivity for MIC-2 in tumor cells, thus final diagnosis of Ewing’s sarcoma was made. The patient was kept for follow up for 3 months and had no symptoms.

Loss of intercellular adhesion leads to differential accumulation of hypericin in bladder cancer

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Lucky, S. Sasidharan; Bhuvaneswari, Ramaswamy; Chin, William W. L.; Lau, Weber K. O.; Olivo, Malini C. D.

2009-06-01

Photodynamic diagnosis (PDD) exploits the photoactive nature of certain compounds, namely photosensitizers, in order to enhance the visual demarcation between normal and neoplastic tissue. Hypericin is one such potent photosensitizer that preferentially accumulate in neoplastic tissue, and fluoresce in the visible spectrum when illuminated with light of an appropriate wavelength. In our study, we investigated the role of E-cadherin in the selective permeation of hypericin in bladder cancer tissues. Clinical studies were done on a series of 43 histologically graded bladder cancer biopsy specimens, obtained from 28 patients who received intravesical instillations with 8μM hypericin solution for at least 2 hours. Immunohistochemical staining was used to assess the expression of E-cadherin, in the cryosectioned tissues. Hypericin uptake was examined by fluorescence microscopy. Immunohistochemical staining showed a clear expression of E-cadherin along the urothelial lining of the normal and pre-malignant tissues. Partial expression of these cell adhesion molecules were still observed in malignant tissues, however there was a loss of expression to variable extends along the urothelium. Thus, loss of intercellular adhesion can be associated with enhanced hypericin permeation through paracellular diffusion.

Toluidine Blue and Hematoxylin and Eosin Stains are Comparable in Evaluating Squamous Cell Carcinoma During Mohs.

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Styperek, Andrew R; Goldberg, Leonard H; Goldschmidt, Laura E; Kimyai-Asadi, Arash

2016-11-01

Histologic examination of tissue is the foundation of Mohs micrographic surgery because determination of surgical margins influences whether additional tissue will be taken. Currently, there is no large focused study comparing toluidine blue (TB) and hematoxylin and eosin (H&E) stains in the evaluation of squamous cell carcinoma (SCC). This study evaluates whether TB and H&E are comparable in assessing the presence of tumor in frozen sections of SCC. One hundred eighty-six randomized slides representing 93 tissue pieces from 36 tumors were examined by 3 Mohs surgeons (1 Accreditation Council for Graduate Medical Education fellow and 2 fellowship-trained surgeons) and compared using a template that documented the presence and location of tumor on the slides. The evaluation of SCC with H&E and TB stains was highly concordant, with concordant identification of SCC in 96%, 96%, and 94% of tissue layers among the 3 Mohs surgeons ARS, LHG, and AK-A, respectively. Toluidine blue and H&E stains are statistically similar in their ability to detect SCC and guide Mohs surgical decision-making.

Liquid dish washing soap: An excellent substitute for xylene and alcohol in hematoxylin and eosin staining procedure

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Surekha Ramulu

2012-01-01

Full Text Available Aims: Liquid dish washing solution (DWS was used as a substitute for xylene to dewax tissue sections during hematoxylin and eosin (H and E staining. The aim was to test and compare the hypothesis that xylene-ethanol free (XEF sections deparaffinized with diluted DWS are better than or at par with the conventional H and E sections. Materials and Methods: Fifty paraffin-embedded tissue blocks was included. One section was stained with conventional HandE (group A and the other with XEF HandE (group B staining method. Slides were scored for parameters: nuclear, cytoplasmic, clarity, uniformity, and crispness of staining. Z test was used for statistical analysis. For accuracy of diagnosis, sensitivity, specificity, positive predictive value, and negative predictive value were tested. Results: Adequate nuclear staining was noted in 94% in group A and 96% in group B, -adequate cytoplasmic staining in 92% in group A and 86% in group B, clarity in 94% of group A and 96% of group B sections, uniform staining in 92% of group A and 80% of group B sections, crisp stain in 96% of group A and 88% of group B sections, and 94% of group A sections stained adequately for diagnosis as compared with 90% in group B sections. Conclusion: Liquid DWS can be used as an alternative and effective substitute to xylene and ethanol in routine HandE staining procedure.

Skin equivalent tissue-engineered construct: co-cultured fibroblasts/ keratinocytes on 3D matrices of sericin hope cocoons.

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Sunita Nayak

Full Text Available The development of effective and alternative tissue-engineered skin replacements to autografts, allografts and xenografts has became a clinical requirement due to the problems related to source of donor tissue and the perceived risk of disease transmission. In the present study 3D tissue engineered construct of sericin is developed using co-culture of keratinocytes on the upper surface of the fabricated matrices and with fibroblasts on lower surface. Sericin is obtained from "Sericin Hope" silkworm of Bombyx mori mutant and is extracted from cocoons by autoclave. Porous sericin matrices are prepared by freeze dried method using genipin as crosslinker. The matrices are characterized biochemically and biophysically. The cell proliferation and viability of co-cultured fibroblasts and keratinocytes on matrices for at least 28 days are observed by live/dead assay, Alamar blue assay, and by dual fluorescent staining. The growth of the fibroblasts and keratinocytes in co-culture is correlated with the expression level of TGF-β, b-FGF and IL-8 in the cultured supernatants by enzyme-linked immunosorbent assay. The histological analysis further demonstrates a multi-layered stratified epidermal layer of uninhibited keratinocytes in co-cultured constructs. Presence of involucrin, collagen IV and the fibroblast surface protein in immuno-histochemical stained sections of co-cultured matrices indicates the significance of paracrine signaling between keratinocytes and fibroblasts in the expression of extracellular matrix protein for dermal repair. No significant amount of pro inflammatory cytokines (TNF-α, IL-1β and nitric oxide production are evidenced when macrophages grown on the sericin matrices. The results all together depict the potentiality of sericin 3D matrices as skin equivalent tissue engineered construct in wound repair.

East coast fever caused by Theileria parva is characterized by macrophage activation associated with vasculitis and respiratory failure

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Respiratory failure and death in East Coast Fever (ECF), a clinical syndrome of African cattle caused by the apicomplexan parasite Theileria parva, has historically been attributed to pulmonary infiltration by infected lymphocytes. However, immunohistochemical staining of tissue from T. parva infect...

Oil Red O and Hematoxylin and Eosin Staining for Quantification of Atherosclerosis Burden in Mouse Aorta and Aortic Root.

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Andrés-Manzano, M Jesús; Andrés, Vicente; Dorado, Beatriz

2015-01-01

Methods for staining tissues with Oil Red O and hematoxylin-eosin are classical histological techniques that are widely used to quantify atherosclerotic burden in mouse tissues because of their ease of use, reliability, and the large amount of information they provide. These stains can provide quantitative data about the impact of a genetic or environmental factor on atherosclerotic burden and on the initiation, progression, or regression of the disease, and can also be used to evaluate the efficacy of drugs designed to prevent or treat atherosclerosis. This chapter provides protocols for quantifying atherosclerotic burden in mouse aorta and aortic root, including methods for dissection, Oil Red O staining, hematoxylin-eosin staining, and image analysis.

Characterization of atherosclerosis by histochemical and immunohistochemical methods in African grey parrots (Psittacus erithacus) and Amazon parrots (Amazona spp.).

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Fricke, Cornelia; Schmidt, Volker; Cramer, Kerstin; Krautwald-Junghanns, Maria-Elisabeth; Dorrestein, Gerry M

2009-09-01

The aim of the study was to characterize atherosclerotic changes in African grey parrots (Psittacus erithacus) and Amazon parrots (Amazona spp.) by histochemical and immunohistochemical methods. Samples of the aorta ascendens and trunci brachiocephalici from 62 African grey parrots and 35 Amazon parrots were stained by hematoxylin and eosin and Elastica van Gieson for grading of atherosclerosis in these birds. Four different stages were differentiated. The incidence of atherosclerosis in the examined parrots was 91.9% in African grey parrots and 91.4% in Amazon parrots. To evaluate the pathogenesis in birds, immunohistochemical methods were performed to demonstrate lymphocytes, macrophages, smooth muscle cells, and chondroitin sulfate. According to the missing lymphocytes and macrophages and the absence of invasion and proliferation of smooth muscle cells in each atherosclerotic stage, "response-to-injury hypothesis" seems inapplicable in parrots. Additionally, we found alterations of vitally important organs (heart, lungs) significantly correlated with atherosclerosis of the aorta ascendens.

Protein tyrosine nitration in chronic intramuscular parasitism: immunohistochemical evaluation of relationships between nitration, and fiber type-specific responses to infection

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Ted H. Elsasser

2011-05-01

Full Text Available The present study was conducted to determine whether preferential muscle catabolism [psoas major (PM > rectus femoris (RF] observed during the chronic intramuscular stage of Sarcocystis cruzi infection could be associated with the pathological consequences of increased protein tyrosine nitration in fibers characteristically more metabolically active due to higher mitochondrial density. Holstein calves were assigned to control (C, or S. cruzi-infected (I groups, n=5/group. Calves were euthanized on day 63 of infection. Samples of RF and PM were prepared for metabolic fiber typing (MFT: slow oxidative, SO – Type I; fast oxidative glycolytic, FOG - Type IIa; fast glycolytic, FG – Type IIb, fiber area, and immunohistochemical localization of fast myosin heavy chain 2a and 2b, nitrotyrosine (NT, and mitochondrial Complex V ATP-synthase. MFT analysis documented that PM contained twice the number of SO fibers compared to RF (32 v 16%, P<0.002. SO and FOG fibers (Both higher in mitochondrial density than FG fibers in both PM and RF were significantly smaller in area in I calves with mean FG areas not different between C and I. Muscle NT content (Western blot of myofibrillar protein fraction increased with infection; NT was immunohistochemically localized into three distinct patterns in fibers: i sparse fiber staining, ii dense punctuate intrafiber staining, and iii pericystic staining. By image analysis, the greatest punctuate intrafiber pixel density of NT was associated with SO fibers from I calves with the NT colocalizing with mitochondrial Complex V – F1F0 ATP synthase. More fibers were positive for the colocalization in PM than RF (P<0.04. The data are consistent with the concept that fibers rich in mitochondria possessing more inherent oxidative energy capacity generate more nitrated proteins than glycolytic fibers and as such are more affected by the proinflammatory response to infections like Sarcocystosis.

Effect of high-frequency near-infrared diode laser irradiation on periodontal tissues during experimental tooth movement in rats.

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Gunji, Hidemi; Kunimatsu, Ryo; Tsuka, Yuji; Yoshimi, Yuki; Sumi, Keisuke; Awada, Tetsuya; Nakajima, Kengo; Kimura, Aya; Hiraki, Tomoka; Hirose, Naoto; Yanoshita, Makoto; Tanimoto, Kotaro

2018-02-05

Tooth movement during orthodontic treatment is associated with bone neoplasticity and bone resorption on the tension and pressure sides. Previous clinical studies have suggested that low-power laser irradiation can accelerate tooth movement during orthodontic treatment, although the underlying mechanism remains unclear. In this study, we used a high-frequency near-infrared diode laser that generates less heat and examined the histologic changes in periodontal tissue during experimental tooth movement with laser irradiation. A nickel-titanium closed coil was mounted between the maxillary left side first molar and incisor of rats to model experimental tooth movement. The laser-irradiation and the control groups were set, and the amount of movement of the first molar on 7th and 14th days after the start of pulling of the first molar tooth on the maxillary left was measured by three-dimensional analysis of µCT. After tooth movement, tissue samples from the mesial and tension sides were collected, and successive horizontal sections were prepared and examined using hematoxylin-eosin and TRAP staining and immunohistochemical staining for RANKL, OPG, ALP, and proliferating cell nuclear antigen (PCNA). Changes in tissue temperature following laser irradiation were also examined. Laser irradiation significantly increased tooth movement compared with non-irradiated controls. Histologic staining of the pressure-side mesial root in laser-irradiated rats revealed enhanced RANKL expression and increased numbers of TRAP-positive cells compared with controls. By contrast, on the tension side, laser irradiation led to increased expression of ALP and PCNA. These data indicate that high-frequency near-infrared diode laser irradiation on the pressure side upregulates RANKL expression and accelerates osteoclast differentiation, facilitating bone resorption, whereas bone formation is induced on the tension side. This study demonstrates that high-frequency near-infrared diode laser

Maintenance of the Extracellular Matrix in Rat Anterior Pituitary Gland: Identification of Cells Expressing Tissue Inhibitors of Metalloproteinases.

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Azuma, Morio; Tofrizal, Alimuddin; Maliza, Rita; Batchuluun, Khongorzul; Ramadhani, Dini; Syaidah, Rahimi; Tsukada, Takehiro; Fujiwara, Ken; Kikuchi, Motoshi; Horiguchi, Kotaro; Yashiro, Takashi

2015-12-25

The extracellular matrix (ECM) is important in creating cellular environments in tissues. Recent studies have demonstrated that ECM components are localized in anterior pituitary cells and affect cell activity. Thus, clarifying the mechanism responsible for ECM maintenance would improve understanding of gland function. Tissue inhibitors of metalloproteinases (TIMPs) are endogenous inhibitors of matrix metalloproteinases and participate in ECM degradation. In this study, we investigated whether cells expressing TIMPs are present in rat anterior pituitary gland. Reverse transcription polymerase chain reaction was used to analyze expression of the TIMP family (TIMP1-4), and cells producing TIMPs in the gland were identified by using in situ hybridization. Expression of TIMP1, TIMP2, and TIMP3 mRNAs was detected, and the TIMP-expressing cells were located in the gland. The TIMP-expressing cells were also investigated by means of double-staining with in situ hybridization and immunohistochemical techniques. Double-staining revealed that TIMP1 mRNA was expressed in folliculostellate cells. TIMP2 mRNA was detected in folliculostellate cells, prolactin cells, and thyroid-stimulating hormone cells. TIMP3 mRNA was identified in endothelial cells, pericytes, novel desmin-immunopositive perivascular cells, and folliculostellate cells. These findings indicate that TIMP1-, TIMP2-, and TIMP3-expressing cells are present in rat anterior pituitary gland and that they are involved in maintaining ECM components.

Maintenance of the Extracellular Matrix in Rat Anterior Pituitary Gland: Identification of Cells Expressing Tissue Inhibitors of Metalloproteinases

International Nuclear Information System (INIS)

Azuma, Morio; Tofrizal, Alimuddin; Maliza, Rita; Batchuluun, Khongorzul; Ramadhani, Dini; Syaidah, Rahimi; Tsukada, Takehiro; Fujiwara, Ken; Kikuchi, Motoshi; Horiguchi, Kotaro; Yashiro, Takashi

2015-01-01

The extracellular matrix (ECM) is important in creating cellular environments in tissues. Recent studies have demonstrated that ECM components are localized in anterior pituitary cells and affect cell activity. Thus, clarifying the mechanism responsible for ECM maintenance would improve understanding of gland function. Tissue inhibitors of metalloproteinases (TIMPs) are endogenous inhibitors of matrix metalloproteinases and participate in ECM degradation. In this study, we investigated whether cells expressing TIMPs are present in rat anterior pituitary gland. Reverse transcription polymerase chain reaction was used to analyze expression of the TIMP family (TIMP1-4), and cells producing TIMPs in the gland were identified by using in situ hybridization. Expression of TIMP1, TIMP2, and TIMP3 mRNAs was detected, and the TIMP-expressing cells were located in the gland. The TIMP-expressing cells were also investigated by means of double-staining with in situ hybridization and immunohistochemical techniques. Double-staining revealed that TIMP1 mRNA was expressed in folliculostellate cells. TIMP2 mRNA was detected in folliculostellate cells, prolactin cells, and thyroid-stimulating hormone cells. TIMP3 mRNA was identified in endothelial cells, pericytes, novel desmin-immunopositive perivascular cells, and folliculostellate cells. These findings indicate that TIMP1-, TIMP2-, and TIMP3-expressing cells are present in rat anterior pituitary gland and that they are involved in maintaining ECM components

Retinoid X receptor gene expression and protein content in tissues of the rock shell Thais clavigera

Energy Technology Data Exchange (ETDEWEB)

Horiguchi, Toshihiro [Research Center for Environmental Risk, National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba, Ibaraki 305-8506 (Japan)], E-mail: thorigu@nies.go.jp; Nishikawa, Tomohiro [Research Center for Environmental Risk, National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba, Ibaraki 305-8506 (Japan); Ohta, Yasuhiko [Department of Veterinary Science, Faculty of Agriculture, Tottori University, 4-101 Koyamacho-Minami, Tottori 680-8553 (Japan); Shiraishi, Hiroaki; Morita, Masatoshi [Research Center for Environmental Risk, National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba, Ibaraki 305-8506 (Japan)

2007-10-15

To elucidate the role of retinoid X receptor (RXR) in the development of imposex caused by organotin compounds in gastropod molluscs, we investigated RXR gene expression and RXR protein content in various tissues of male and female wild rock shells (Thais clavigera). Quantitative real-time polymerase chain reaction, Western blotting, and immunohistochemistry with a commercial antibody against human RXR {alpha} revealed that RXR gene expression was significantly higher in the penises of males and imposex-exhibiting females than in the penis-forming areas of normal females (P < 0.01 and P < 0.05, respectively). Western blotting demonstrated that the antibody could detect rock shell RXR and showed that the male penis had the highest content of RXR protein among the analyzed tissues of males and normal females. Immunohistochemical staining revealed nuclear localization of RXR protein in the epithelial and smooth muscle cells of the vas deferens and in the interstitial or connective tissues and epidermis of the penis in males and imposex-exhibiting females. RXR could be involved in the mechanism of induction of male-type genitalia (penis and vas deferens) by organotin compounds in female rock shells.

Interstitial mononuclear cell infiltrates in chronic rejection of the kidney and correlation with peripheral blood.

OpenAIRE

Jeong, H. J.; Hong, S. W.; Kim, Y. S.; Kim, M. S.; Choi, I. H.; Park, K.; Choi, I. J.

1996-01-01

To investigate the characteristics of interstitial inflammatory cells and possible involvement of nudelta T cells, 16 renal allograft biopsies showing chronic rejection were stained by immunohistochemical method and correlated with the data of peripheral blood evaluated by flow cytometry. For immunophenotyping, fresh frozen sections were stained with monoclonal antibodies against CD3, CD4, CD8, CD68, CD56, TCRdelta1 and HLA DR. Paraffin embedded tissue was stained with CD45RO, CD20-Cy and CD6...

Richly innervated soft tissues covering the superficial aspect of the extensor origin in patients with chronic painful tennis elbow - Implication for treatment?

Science.gov (United States)

Spang, C; Alfredson, H

2017-06-01

Tennis elbow is difficult to treat. The results of surgical treatments are not convincing. Treatment studies on Achilles and patellar tendinopathy targeting the richly innervated and vascularized soft tissues outside the tendon have shown promising outcomes. The innervation patterns in the fibrous/fatty tissues superficially to the elbow extensor origin have not been clarified. Nine tissue specimens from the fibrous/fatty tissue covering the extensor origin was taken from seven patients (mean age: 45 years) undergoing surgical treatment for chronic painful tennis elbow. The specimens were stained for morphology (haematoxylin and eosin, H and E) and immunohistochemically for general nerve marker protein gene product 9.5 (PGP 9.5) and markers for sympathetic (tyrosine hydroxylase, TH) and sensory nerve fibres (calcitonin gene-related peptide, CGRP). All specimens contained multiple blood vessels and nerve structures indicated by morphology and immunoreactions. There was a frequent occurrence of TH reactions, especially peri-vascularly, but also in nerve fascicles. Immunoreactions for CGRP were seen in nerve fascicles and isolated nerve fibres. The results provide new information on the innervation patterns of the superficial tissues of the extensor origin and their potential as source of tennis elbow pain. IV.

Rapid in vivo vertical tissue sectioning by multiphoton tomography

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Batista, Ana; Breunig, Hans Georg; König, Karsten

2018-02-01

A conventional tool in the pathological field is histology which involves the analysis of thin sections of tissue in which specific cellular structures are stained with different dyes. The process to obtain these stained tissue sections is time consuming and invasive as it requires tissue removal, fixation, sectioning, and staining. Moreover, imaging of live tissue is not possible. We demonstrate that multiphoton tomography can provide within seconds, non-invasive, label-free, vertical images of live tissue which are in quality similar to conventional light micrographs of histologic stained specimen. In contrast to conventional setups based on laser scanning which image horizontally sections, the vertical in vivo images are directly recorded by combined line scanning and timed adjustments of the height of the focusing optics. In addition, multiphoton tomography provides autofluorescence lifetimes which can be used to determine the metabolic states of cells.

Host inflammatory response to polypropylene implants: insights from a quantitative immunohistochemical and birefringence analysis in a rat subcutaneous model

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Alessandro Prudente

2016-06-01

Full Text Available ABSTRACT Objectives To describe acute and sub acute aspects of histological and immunohistochemical response to PP implant in a rat subcutaneous model based on objective methods. Materials and Methods Thirty rats had a PP mesh subcutaneously implanted and the same dissection on the other side of abdomen but without mesh (sham. The animals were euthanized after 4 and 30 days. Six slides were prepared using the tissue removed: one stained with hematoxylin-eosin (inflammation assessment; one unstained (birefringence evaluation and four slides for immunohistochemical processing: IL-1 and TNF-α (pro-inflammatory cytokines, MMP-2 (collagen metabolism and CD-31 (angiogenesis. The area of inflammation, the birefringence index, the area of immunoreactivity and the number of vessels were objectively measured. Results A larger area of inflammatory reaction was observed in PP compared to sham on the 4th and on the 30th day (p=0.0002. After 4 days, PP presented higher TNF (p=0.0001 immunoreactivity than sham and no differences were observed in MMP-2 (p=0.06 and IL-1 (p=0.08. After 30 days, a reduction of IL-1 (p=0.010 and TNF (p=0.016 for PP and of IL-1 (p=0.010 for sham were observed. Moreover, area of MMP-2 immunoreactivity decreased over time for PP group (p=0.018. Birefringence index and vessel counting showed no differences between PP and sham (p=0.27 and p=0.58, respectively. Conclusions The implantation of monofilament and macroporous polypropylene in the subcutaneous of rats resulted in increased inflammatory activity and higher TNF production in the early post implant phase. After 30 days, PP has similar cytokines immunoreactivity, vessel density and extracellular matrix organization.

Nonlinear multicontrast microscopy of hematoxylin-and-eosin-stained histological sections

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Tuer, Adam; Tokarz, Danielle; Prent, Nicole; Cisek, Richard; Alami, Jennifer; Dumont, Daniel J.; Bakueva, Ludmila; Rowlands, John; Barzda, Virginijus

2010-03-01

Imaging hematoxylin-and-eosin-stained cancerous histological sections with multicontrast nonlinear excitation fluorescence, second- and third-harmonic generation (THG) microscopy reveals cellular structures with extremely high image contrast. Absorption and fluorescence spectroscopy together with second hyperpolarizability measurements of the dyes shows that strong THG appears due to neutral hemalum aggregation and is subsequently enhanced by interaction with eosin. Additionally, fluorescence lifetime imaging microscopy reveals eosin fluorescence quenching by hemalums, showing better suitability of only eosin staining for fluorescence microscopy. Multicontrast nonlinear microscopy has the potential to differentiate between cancerous and healthy tissue at a single cell level.

An additive manufacturing-based PCL-alginate-chondrocyte bioprinted scaffold for cartilage tissue engineering.

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Kundu, Joydip; Shim, Jin-Hyung; Jang, Jinah; Kim, Sung-Won; Cho, Dong-Woo

2015-11-01

Regenerative medicine is targeted to improve, restore or replace damaged tissues or organs using a combination of cells, materials and growth factors. Both tissue engineering and developmental biology currently deal with the process of tissue self-assembly and extracellular matrix (ECM) deposition. In this investigation, additive manufacturing (AM) with a multihead deposition system (MHDS) was used to fabricate three-dimensional (3D) cell-printed scaffolds using layer-by-layer (LBL) deposition of polycaprolactone (PCL) and chondrocyte cell-encapsulated alginate hydrogel. Appropriate cell dispensing conditions and optimum alginate concentrations for maintaining cell viability were determined. In vitro cell-based biochemical assays were performed to determine glycosaminoglycans (GAGs), DNA and total collagen contents from different PCL-alginate gel constructs. PCL-alginate gels containing transforming growth factor-β (TGFβ) showed higher ECM formation. The 3D cell-printed scaffolds of PCL-alginate gel were implanted in the dorsal subcutaneous spaces of female nude mice. Histochemical [Alcian blue and haematoxylin and eosin (H&E) staining] and immunohistochemical (type II collagen) analyses of the retrieved implants after 4 weeks revealed enhanced cartilage tissue and type II collagen fibril formation in the PCL-alginate gel (+TGFβ) hybrid scaffold. In conclusion, we present an innovative cell-printed scaffold for cartilage regeneration fabricated by an advanced bioprinting technology. Copyright © 2013 John Wiley & Sons, Ltd.

Immunohistochemical differentiation of high-grade prostate carcinoma from urothelial carcinoma.

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Chuang, Ai-Ying; DeMarzo, Angelo M; Veltri, Robert W; Sharma, Rajni B; Bieberich, Charles J; Epstein, Jonathan I

2007-08-01

The histologic distinction between high-grade prostate cancer and infiltrating high-grade urothelial cancer may be difficult, and has significant implications because each disease may be treated very differently (ie, hormone therapy for prostate cancer and chemotherapy for urothelial cancer). Immunohistochemistry of novel and established prostatic and urothelial markers using tissue microarrays (TMAs) were studied. Prostatic markers studied included: prostate-specific antigen (PSA), prostein (P501s), prostate-specific membrane antigen (PSMA), NKX3.1 (an androgen-related tumor suppressor gene), and proPSA (pPSA) (precursor form of PSA). "Urothelial markers" included high molecular weight cytokeratin (HMWCK), p63, thrombomodulin, and S100P (placental S100). TMAs contained 38 poorly differentiated prostate cancers [Gleason score 8 (n=2), Gleason score 9 (n=18), Gleason score 10 (n=18)] and 35 high-grade invasive urothelial carcinomas from radical prostatectomy and cystectomy specimens, respectively. Each case had 2 to 8 tissue spots (0.6-mm diameter). If all spots for a case showed negative staining, the case was called negative. The sensitivities for labeling prostate cancers were PSA (97.4%), P501S (100%), PSMA (92.1%), NKX3.1 (94.7%), and pPSA (94.7%). Because of PSA's high sensitivity on the TMA, we chose 41 additional poorly differentiated primary (N=36) and metastatic (N=5) prostate carcinomas which showed variable PSA staining at the time of diagnosis and performed immunohistochemistry on routine tissue sections. Compared to PSA, which on average showed 18.8% of cells with moderate to strong positivity, cases stained for P501S, PSMA, and NKX3.1 had on average 42.5%, 53.7%, 52.9% immunoreactivity, respectively. All prostatic markers showed excellent specificity. HMWCK, p63, thrombomodulin, and S100P showed lower sensitivities in labeling high-grade invasive urothelial cancer in the TMAs with 91.4%, 82.9%, 68.6%, and 71.4% staining, respectively. These urothelial

Immunolocalisation of oestrogen receptor beta in human tissues.

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Taylor, A H; Al-Azzawi, F

2000-02-01

Oestrogens exert their actions via specific nuclear protein receptors that are members of the steroid/thyroid receptor superfamily of transcription factors. Recently, a second oestrogen receptor (ERbeta) has been cloned, and using reverse transcription-PCR and immunohistochemistry it has been shown to have a wide tissue distribution in the rat that is distinct from the classical oestrogen receptor, ERalpha. Using commercial polyclonal antisera against peptides specific to human ERbeta, we have determined the sites of ERbeta expression in archival and formalin-fixed human tissue and compared its expression with that of ERalpha. ERbeta was localised to the cell nuclei of a wide range of normal adult human tissues including ovary, Fallopian tube, uterus, lung, kidney, brain, heart, prostate and testis. In the ovary, ERbeta was present in multiple cell types including granulosa cells in small, medium and large follicles, theca and corpora lutea, whereas ERalpha was weakly expressed in the nuclei of granulosa cells, but not in the theca nor in the copora lutea. In the endometrium, both ERalpha and ERbeta were observed in luminal epithelial cells and in the nuclei of stromal cells but, significantly, ERbeta was weak or absent from endometrial glandular epithelia. Epithelial cells in most male tissues including the prostate, the urothelium and muscle layers of the bladder, and Sertoli cells in the testis, were also immunopositive for ERbeta. Significant ERbeta immunoreactivity was detected in most areas of the brain, with the exception of the hippocampus - a tissue that stained positively for ERalpha. In conclusion, the almost ubiquitous immunohistochemical localisation of ERbeta indicates that ERbeta may play a major role in the mediation of oestrogen action. The differential expression of ERalpha and ERbeta in some of these tissues suggests a more complex control mechanism in oestrogenic potential than originally envisioned.

The Role of Immunohistochemical Biomarkers as Prognostic Factors by the Use of a Tissue Microarray in Breast Cancer Patients Under 45-years-old

International Nuclear Information System (INIS)

Kim, Eun Seog; Choi, Doo Ho; Jin, So Young

2008-01-01

This study evaluates the association of estrogen receptor (ER), progesterone receptor (PR), Her-2, COX-2, and survivin with the clinicopathological features and outcomes in young Korean women with breast cancer using recently developed tissue microarray (TMA) technology. A cohort of 212 young patients with breast cancer diagnosed at the age of 45 years or younger from March 1994 to August 2005, were enrolled in this study. The age range of patients was 23∼45 years (median age, 39 years). The minimum and median follow-up periods were 24 months and 60 months, respectively. Serial sections of primary tumors were processed by the use of a TMA for immunohistochemical staining for five biomarkers. The correlation of these five biomarkers and the clinicopathological features and outcomes were analyzed by statistical methods. The majority of the patients were stage T1 (90 patients) or T2 (101 patients), and 105 patients (49.5%) had an axillary node metastasis. The 5-year overall and relapse free survival rates for all of the patients were 90.4% and 82.3%, respectively, and 36 patients had a locoregional or distant metastasis as a first event. Positive expression of ER, PR, Her-2, COX-2, and survivin was determined in 38.2%, 45.3%, 25.9%, 41.5%, and 43.4%, of the tumor samples, respectively. Tumor stage, nodal status, age, as well as expression of ER, PR, and HER-2 status were significantly associated with the disease free survival rate. Tumor stage, nodal status, as well as expression of ER, PR, and HER-2 were significantly related with the overall survival rate. Expression of COX-2 and survivin were not single independent prognostic factors for the disease free and overall survival rate although co-expression of HER-2 and COX-2 had a tendency as a poor prognostic factor. By multivariate analysis, only T stage and lymph node status were significant prognostic factors, and ER status was a marginally significant prognostic factor (p=0.075). Expression of ER, PR and HER-2

Immunohistochemical study of DNA topoisomerase I, DNA topoisomerase II alpha, p53, and Ki-67 in oral preneoplastic lesions and oral squamous cell carcinomas.

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Hafian, Hilal; Venteo, Lydie; Sukhanova, Alyona; Nabiev, Igor; Lefevre, Benoît; Pluot, Michel

2004-06-01

Human DNA topoisomerase I (topo I) is the molecular target of the camptothecin group of anticancer drugs. Laboratory studies have shown that the cellular response to topo I-targeted drugs depends on the topo I expression and DNA replication rate and the apoptotic pathway activity. In this study, we tested potential indicators of the sensitivity of topo I-targeted drugs in 36 cases of oral squamous cell carcinoma (OSCC). Formalin-fixed, paraffin-embedded tissue sections were immunostained with monoclonal antibodies against Ki-67, p53, and topo I, and with polyclonal antibodies against DNA topoisomerase II-alpha (topo II-alpha). These markers were also tested in 18 epithelial hyperplastic lesions and 18 mild dysplasias. Immunostaining was quantified by the percentage of stained nuclei in each sample (the labeling index); 200 immunoreactive epithelial nuclei were counted per case for each antibody. The results support the possibility of using topo II-alpha staining for assessing the proliferative activity. High expression of topo II-alpha and topo I in OSCCs suggests that they may serve as potential indicators of sensitivity to topo I inhibitors. However, the apoptotic pathway assessed by p53 immunostaining was found to be uninformative. Analysis of the relationship between immunohistochemical results and clinical and pathologic parameters (the T and N stages and differentiation) showed that only the differentiation parameter correlated with the topo I expression rate. Thus, significant increase in the topo I expression in the poorly differentiated OSCCs suggests their higher sensitivity to drug treatment.

Metastatic hepatocellular carcinoma to the skin staining positive with HMB-45.

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Gross, Joshua A; Perniciaro, Charles; Gross, David J; Barksdale, Sarah K

2012-02-01

Hepatocellular carcinoma (HCC) is uncommonly observed as a cutaneous metastasis. We report a 76-year-old man with metastatic HCC to the skin of the nasal ala, diagnosed antecedent to the primary tumor. HCC was confirmed by positive immunostaining with Hep Par 1 in tissue from the metastasis and from a needle biopsy of the primary lesion. In addition, tumor cells from both the metastasis and liver stained positive with HMB-45. To our knowledge, HMB-45 positive staining has not been reported in either primary or metastatic HCC.

The effects of antidepressants and pilocarpine on rat parotid glands: an immunohistochemical study

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Tatiana Maria Folador Mattioli

2011-01-01

Full Text Available OBJECTIVES: To evaluate the effects of antidepressants and pilocarpine on the quantity of myoepithelial cells and on the proliferation index of the epithelial cells of rat parotid glands. INTRODUCTION: Hyposalivation, xerostomia, and alterations in saliva composition are important clinical side effects related to the use of antidepressants. METHODS: Ninety male Wistar rats were allocated to nine groups. The control groups received saline for 30 (group C30 or 60 days (group C60 or pilocarpine for 60 days (group Pilo. The experimental groups were administered fluoxetine (group F30 or venlafaxine for 30 days (group V30; fluoxetine (group FS60 or venlafaxine (group VS60 with saline for 60 days; or fluoxetine (group FP60 or venlafaxine (group VP60 with pilocarpine for 60 days. Parotid gland specimens were processed, and the immunohistochemical expression of calponin and proliferating cell nuclear anti-antigen on the myoepithelial and parenchymal cells, respectively, was evaluated. Analysis of variance (ANOVA, Tukey HSD and Games-Howell tests were applied to detect differences among groups (p<0.05. RESULTS: Compared with the controls, chronic exposure to antidepressants was associated with an increase in the number of positively stained cells for calponin. In addition, venlafaxine administration for 30 days was associated with an increase in the number of positively stained cells for proliferating cell nuclear anti-antigen. Fluoxetine and pilocarpine (group FP60 induced a significant decrease in the number of positively stained cells for calponin compared with all other groups. CONCLUSIONS: The number of positively stained cells for calponin increased after chronic administration of antidepressants. The proliferation index of the epithelial cells of rat parotid glands was not altered by the use of antidepressants for 60 days.

Immunohistochemical study of macrophage migration inhibitory factor in rat liver fibrosis induced by thioacetamide

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Y Hori

2009-06-01

Full Text Available Macrophage migration inhibitory factor (MIF is a molecule known to regulate macrophage accumulation at sites of inflammation. To elucidate the role of MIF in progression of liver fibrosis, the immunohistochemical localization of MIF and macrophages in the liver were examined. Male Wistar rats received thioacetamide (TA injections (200 mg/kg, i.p. for 1 or 6 weeks. In biochemical and histological tests, it was confirmed that liver fibrosis was induced. In immunohistochemical analyses, the expression of MIF protein was seen in hepatocytes in the areas extending out from the central veins to the portal tracts. In particular, at 6 weeks, immunoreactivity was detected in degenerated hepatocytes adjacent to the fibrotic areas but hardly observed in the fibrotic areas. On the other hand, a number of exudate macrophages stained by antibody ED1 were seen in the areas from the central veins to the portal tracts at 1 week and in the fibrotic areas at 6 weeks. Macrophages also showed a significant increase in number as compared with controls. These results revealed that there was a close relationship between the appearance of MIF expression and ED1-positive exudate macrophages in degenerated hepatocytes during the progression of TA-induced liver fibrosis.

Immunohistochemical study of macrophage migration inhibitory factor in rat liver fibrosis induced by thioacetamide.

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Hori, Y; Sato, S; Yamate, J; Kurasaki, M; Nishihira, J; Hosokawa, T; Fujita, H; Saito, T

2003-01-01

Macrophage migration inhibitory factor (MIF) is a molecule known to regulate macrophage accumulation at sites of inflammation. To elucidate the role of MIF in progression of liver fibrosis, the immunohistochemical localization of MIF and macrophages in the liver were examined. Male Wistar rats received thioacetamide (TA) injections (200 mg/kg, i.p.) for 1 or 6 weeks. In biochemical and histological tests, it was confirmed that liver fibrosis was induced. In immunohistochemical analyses, the expression of MIF protein was seen in hepatocytes in the areas extending out from the central veins to the portal tracts. In particular, at 6 weeks, immunoreactivity was detected in degenerated hepatocytes adjacent to the fibrotic areas but hardly observed in the fibrotic areas. On the other hand, a number of exudate macrophages stained by antibody ED1 were seen in the areas from the central veins to the portal tracts at 1 week and in the fibrotic areas at 6 weeks. Macrophages also showed a significant increase in number as compared with controls. These results revealed that there was a close relationship between the appearance of MIF expression and ED1-positive exudate macrophages in degenerated hepatocytes during the progression of TA-induced liver fibrosis.

Survivin Expression in Colorectal Adenocarcinoma Using Tissue Micro array

International Nuclear Information System (INIS)

Abd El-Hamed, A.

2005-01-01

The additional prognostic information closely related to tumor cell biology is essential for the identification of patients with poor prognosis. Survivin, an identified inhibitor of apoptosis, is unique for its expression in human malignancies but not in normal adult cells. This study examined the expression, and potential prognostic value of survivin in colorectal adenocarcinoma (CRC) on tissue micro array (TMA) sections. Analysis of large numbers of tissue samples, improved tissue salvage, cost reduction, ease of interpretation, and significant time saving were realized by using the arrays. Material and Methods: Two-hundred and eighty cases of colorectal adenocarcinoma were arrayed. Immunohistochemical stains of TMA sections were performed for survivin, bcl-2, and p53. Cases were followed up for 5 years. Survivin was detected in 147 of 230 cases (63.9%). No expression of survivin was observed in normal tissues. There was no correlation between survivin immunoreactivity and age, sex, tumor site, tumor size, histopathologic subtype, tumor grade and clinical stage(ρ> 0.05). Prevalence of survivin expression was significantly higher in bcl-2 positive than in bcl-2 negative cases (88.1 % versus 42.1 %, (ρ<0.0001), but was not associated with p53 ((ρ=0.09). The 5-year disease free survival (DFS) for patients with survivin positive colorectal adenocarcinoma was significantly lower than that for patients with survivin negative tumors (46% versus 68.7%, (ρ<0.001). Survivin expression in colorectal adenocarcinoma provides an important prognostic parameter and targeted antagonists of survivin may be beneficial as apoptosis-based therapy for colon cancer

Multiphoton microscopy as a diagnostic imaging modality for pancreatic neoplasms without hematoxylin and eosin stains

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Chen, Youting; Chen, Jing; Chen, Hong; Hong, Zhipeng; Zhu, Xiaoqin; Zhuo, Shuangmu; Chen, Yanling; Chen, Jianxin

2014-09-01

Hematoxylin and eosin (H&E) staining of tissue samples is the standard approach in histopathology for imaging and diagnosing cancer. Recent reports have shown that multiphoton microscopy (MPM) provides better sample interface with single-cell resolution, which enhances traditional H&E staining and offers a powerful diagnostic tool with potential applications in oncology. The purpose of this study was to further expand the versatility of MPM by establishing the optical parameters required for imaging unstained histological sections of pancreatic neoplasms, thereby providing an efficient and environmentally sustainable alternative to H&E staining while improving the accuracy of pancreatic cancer diagnoses. We found that the high-resolution MPM images clearly distinguish between the structure of normal pancreatic tissues compared with pancreatic neoplasms in unstained histological sections, and discernable differences in tissue architecture and cell morphology between normal versus tumorigenic cells led to enhanced optical diagnosis of cancerous tissue. Moreover, quantitative assessment of the cytomorphological features visualized from MPM images showed significant differences in the nuclear-cytoplasmic ratios of pancreatic neoplasms compared with normal pancreas, as well as further distinguished pancreatic malignant tumors from benign tumors. These results indicate that the MPM could potentially serve as an optical tool for the diagnosis of pancreatic neoplasms in unstained histological sections.

Histochemical characterization of human osteochondral tissue: comparison between healthy cartilage, arthrotic tissues, and cartilage defect treated with MACI technique

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F. Tessarolo

2011-01-01

Full Text Available Matrix-induced sutologous chondrocytes implantation (MACI is a promising technique for the treatment of articular cartilage lesions, but long time outcome have to be established. We developed and optimized specific techniques of histochemical staining to characterize healthy and pathologic osteochondral tissue. Seven different staining protocols were applied to assess tissue architecture, cells morphology, proteoglycan content, and collagen fibers distribution. Potentialities of histochemical staining and histomorphology of biopsies from second look arthroscopy will be presented.

Immunohistochemical and morphological features of a small bowel leiomyoma in a black crested macaque (Macaca nigra

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Aristizabal-Arbelaez Mónica

2012-06-01

Full Text Available Abstract Background Spontaneous gastrointestinal neoplasms in non-human primates are commonly seen in aged individuals. Due to genetic similarities between human and non-human primates, scientists have shown increasing interest in terms of comparative oncology studies. Case presentation The present study is related to a case of an intestinal leiomyoma in a black crested macaque (Macaca nigra, kept on captivity by Matecaña Zoo, Pereira City, Colombia. The animal had abdominal distension, anorexia, vomiting, diarrhea and behavioral changes. Clinical examination showed an increased volume in the upper right abdominal quadrant caused by a neoplastic mass. The patient died during the surgical procedure. Necropsy revealed several small nodules in the peritoneum with adhesion to different portions of the small and large intestines, liver, stomach and diaphragm. Tissue samples were collected, routinely processed and stained by H&E. Microscopic examination revealed a mesenchymal tumor limited to tunica muscularis, resembling normal smooth muscle cells. Neoplastic cells were positive for alpha-smooth muscle actin and vimentin, and negative for cytokeratin AE1/AE3 by immunohistochemistry. Those morphological and immunohistochemical findings allowed to diagnose the intestinal leiomyoma referred above. Conclusion Neoplastic diseases in primates have multifaceted causes. Their manifestations are understudied, leading to a greater difficulty in detection and measurement of the real impact provides by this disease.

Amalgam stained dentin: a proper substrate for bonding resin composite?

NARCIS (Netherlands)

Scholtanus, J.D.

2016-01-01

Nowadays the use of dental amalgam is mostly abandoned and substituted by tooth colored resin composites that can be bonded to teeth tissues by adhesive techniques. The aim of this thesis was to find out whether dark stained dentin, as often observed after removal of amalgam restorations and

Tissue concentrations of prostate-specific antigen in prostatic carcinoma and benign prostatic hyperplasia.

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Pretlow, T G; Pretlow, T P; Yang, B; Kaetzel, C S; Delmoro, C M; Kamis, S M; Bodner, D R; Kursh, E; Resnick, M I; Bradley, E L

1991-11-11

Prostate-specific antigen (PSA), as measured in peripheral blood, is currently the most widely used marker for the assessment of tumor burden in the longitudinal study of patients with carcinoma of the prostate (PCA). Studies from other laboratories have led to the conclusion that a given volume of PCA causes a much higher level of PSA in the peripheral circulation of patients than a similar volume of prostate without carcinoma. We have evaluated PSA in the resected tissues immunohistochemically and in extracts of PCA and of prostates resected because of benign prostatic hyperplasia (BPH) with an enzyme-linked immunosorbent assay. Immunohistochemical results were less quantitative than but consistent with the results of the ELISA of tissue extracts. Immunohistochemically, there was considerable heterogeneity in the expression of PSA by both PCA and BPH both within and among prostatic tissues from different patients. While the levels of expression of PSA in these tissues overlap broadly, PSA is expressed at a lower level in PCA than in BPH when PSA is expressed as a function of wet weight of tissue (p = 0.0095), wet weight of tissue/% epithelium (p less than 0.0001), protein extracted from the tissue (p = 0.0039), or protein extracted/% epithelium (p less than 0.0001).

Effects of surface hydrophilicity and microtopography on early stages of soft and hard tissue integration at non-submerged titanium implants: an immunohistochemical study in dogs.

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Schwarz, Frank; Ferrari, Daniel; Herten, Monika; Mihatovic, Ilja; Wieland, Marco; Sager, Martin; Becker, Jürgen

2007-11-01

The aim of the present study was to investigate the effects of surface hydrophilicity and microtopography on soft and hard tissue integration at non-submerged titanium implants. Implantation of conventional sand-blasted large grit and acid-etched (SLA) and chemically modified SLA (modSLA) titanium implants with differently structured transmucosal surfaces (SLA implants: machined [M-SLA] or SLA [SLA-SLA]; modSLA implants: mod acid-etched [modA] [modA-modSLA] or modSLA [modSLA-modSLA]) was performed bilaterally in the upper and lower jaws of 15 beagle dogs. The animals were sacrificed after 1, 4, 7, 14, or 28 days. Tissue reactions were assessed histomorphometrically and immunohistochemically using monoclonal antibodies to transglutaminase II (angiogenesis) and osteocalcin. Although the junctional epithelium commonly was separated from M-SLA and SLA-SLA implants by a gap, the epithelial cells appeared to be in close contact with modA-modSLA surfaces after 14 days of healing. Moreover, modA-modSLA and modSLA-modSLA groups showed a well-vascularized subepithelial connective tissue exhibiting collagen fibers that started to extend and attach partially perpendicular to the implant surface. The highest and statistically significant mean bone-to-implant contact areas were observed in the modA-modSLA and modSLA-modSLA groups at days 7, 14, and 28. Within the limits of this study, it may be concluded that soft and hard tissue integration was influenced mainly by surface hydrophilicity rather than by microtopography.

Immunohistochemical characteristics of neurons in nodose ganglia projecting to the different chambers of the rat heart.

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Kosta, Vana; Guić, Maja Marinović; Aljinović, Jure; Sapunar, Damir; Grković, Ivica

2010-06-24

Despite the contribution of nodose ganglia neurons to the innervation of the heart being the subject of several studies, specific neuronal subpopulations innervating the four different chambers of the heart have not been distinguished. In our study, the application of Fast Blue-loaded patch to the epicardial surface of different chambers of the rat heart (the right or left atrium or the right or left ventricle) resulted in labeling of discrete populations of immunohistochemically diverse neurons. About one half (55%) of these neurons showed immunoreactivity for the 200-kDa neurofilament protein (marker of myelinated neurons), with a higher proportion of positive staining among neurons projecting to the left than to the right ventricle. Isolectin B4 immunoreactivity (characteristic for a subset of nonmyelinated non-peptidergic neurons) was more abundant among neurons projecting to the right side of the heart (right atria and right ventricles) compared to the left side (23% vs. 16%). Calretinin immunoreactivity (possible marker of mechanosensitive neurons) was significantly higher among neurons projecting to the ventricles than among those projecting to atria (36% vs. 11%). These findings reveal that chambers of the rat heart are innervated with immunohistochemically different subpopulations of neurons from the nodose ganglia.

A comparative study to evaluate liquid dish washing soap as an alternative to xylene and alcohol in deparaffinization and hematoxylin and eosin staining.

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Pandey, Pinki; Dixit, Alok; Tanwar, Aparna; Sharma, Anuradha; Mittal, Sanjeev

2014-07-01

Our study presents a new deparaffinizing and hematoxylin and eosin (H and E) staining method that involves the use of easily available, nontoxic and eco-friendly liquid diluted dish washing soap (DWS) by completely eliminating expensive and hazardous xylene and alcohol from deparaffinizing and rehydration prior to staining, staining and from dehydration prior to mounting. The aim was to evaluate and compare the quality of liquid DWS treated xylene and alcohol free (XAF) sections with that of the conventional H and E sections. A total of 100 paraffin embedded tissue blocks from different tissues were included. From each tissue block, one section was stained with conventional H and E (normal sections) and the other with XAF H and E (soapy sections) staining method. Slides were scored using five parameters: Nuclear, cytoplasmic, clarity, uniformity, and crispness of staining. Z-test was used for statistical analysis. Soapy sections scored better for cytoplasmic (90%) and crisp staining (95%) with a statistically significant difference. Whereas for uniformity of staining, normal sections (88%) scored over soapy sections (72%) (Z = 2.82, P 0.05). Liquid DWS is a safe and efficient alternative to xylene and alcohol in deparaffinization and routine H and E staining procedure. We are documenting this project that can be used as a model for other histology laboratories.

Tumour cell expression of C4.4A, a structural homologue of the urokinase receptor, correlates with poor prognosis in non-small cell lung cancer

DEFF Research Database (Denmark)

Hansen, Line V.; Skov, Birgit G; Ploug, Michael

2007-01-01

expression. In the present study, we therefore explored the possible association between C4.4A expression and prognosis in patients with non-small cell lung cancer (NSCLC). EXPERIMENTAL DESIGN: Tissue sections from 108 NSCLC patients were subjected to immunohistochemical staining using a polyclonal antibody...

Urokinase-type plasminogen activator receptor (uPAR) on tumor-associated macrophages is a marker of poor prognosis in colorectal cancer

DEFF Research Database (Denmark)

Illemann, Martin; Laerum, Ole Didrik; Hasselby, Jane Preuss

2014-01-01

Patients were identified from a population-based prospective study of 4990 individuals with symptoms associated with colorectal cancer (CRC). A total of 244 CRC tissue samples were available for immunohistochemical staining of uPAR, semiquantitatively scored at the invasive front, and in the tumo...

Immunohistochemical study of the digestive tract of Oligosarcus hepsetus

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Vieira-Lopes, Danielle A; Pinheiro, Nadja L; Sales, Armando; Ventura, Adriana; Araújo, Francisco G; Gomes, Iracema D; Nascimento, Aparecida A

2013-01-01

AIM: To describe the histology of the digestive tract and to investigate the occurrence of endocrine cells in Oligosarcus hepsetus (O. hepsetus). METHODS: The digestive tract (DT) of O. hepsetus was divided into esophagus, two stomach regions (glandular and non-glandular) and two intestinal regions (anterior and posterior). These specimens were processed by routine histological techniques and stained with hematoxylin-eosin, Gomori’s trichrome, periodic acid Schiff (PAS) and Alcian blue (AB). An immunohistochemical method using avidin-biotin-peroxidase was employed. RESULTS: The esophagus is lined with a non-keratinized stratified squamous epithelium that is reactive to PAS and AB. The stomach has a mucosa lined with a simple columnar epithelium with mucus-secreting cells that are reactive only to PAS. The intestine has a simple columnar epithelium with a brush border and goblet cells that are reactive to PAS and AB. Somatostatin, serotonin and cholecystokinin immunoreactive cells were identified throughout the DT. CONCLUSION: This study revealed adaptations for the species’ diet and showed that the distribution and relative frequency of immunoreactive cells are similar to those of other fish. PMID:23569337

Immunohistochemical Study of p53 Expression in Patients with Erosive and Non-Erosive Oral Lichen Planus

Science.gov (United States)

Shiva, Atena; Zamanian, Ali; Arab, Shahin; Boloki, Mahsa

2018-01-01

Statement of the Problem: Oral lichen planus is a common mucocutaneous lesion with a chronic inflammatory process mediated by immune factors while a few cases of the disease become malignant. Purpose: This study aimed to determine the frequency of p53 marker as a tumor suppressor in patients with erosive and non-erosive oral lichen planus (OLP) by using immunohistochemical methods. Materials and Method: This descriptive cross-sectional study investigated the p53 expression in 16 erosive OLP, 16 non-erosive OLP samples, and 8 samples of normal oral mucosa through immunohistochemistry. The percentage of stained cells in basal and suprabasal layers, and inflammatory infiltrate were graded according to the degree of staining; if 0%, 50% of the cells were stained, they were considered as (-), (+), (++), (+++) and (++++), respectively. The obtained data was statistically analyzed and compared by using Chi square and Fisher’s exact test. Results: The mean percentage of p53 positive cells in erosive OLP (34.5±14.2) was considerably higher than that in non-erosive OLP (23.8±10.4) and normal mucosa (17.5±17). There was a significant difference among the three groups of erosive, non-erosive and control in terms of staining intensity. No significant difference existed between the patients’ age and sex in the two OLP groups. Conclusion: The increased incidence of p53 from normal mucosa to erosive OLP indicated the difference between biological behavior of erosive and non-erosive OLP. It can be claimed that the erosive OLP has great premalignant potential compared with the non-erosive one.

Immunohistochemical expression of vascular endothelial growth factor in keratocystic odontogenic tumor, dentigerous cyst, and radicular cyst: A comparative study.

Science.gov (United States)

Khajuria, Nidhi; Metgud, Rashmi; Naik, Smitha; Lerra, Sahul; Tiwari, Priya; Mamta; Katakwar, Payal; Tak, Anirudh

2016-01-01

Cyst and tumors arise from tissue remains of odontogenesis, these interactions have been considered to play an important role in the tumorigenesis of odontogenic lesions. The connective tissue stroma has an essential role in the preservation of epithelial tissues and minor alterations in the epithelium are followed by corresponding changes in the stroma, such as angiogenesis. Vascular endothelial growth factor (VEGF) is considered the first factor which maintains its position as the most critical driver of vascular formation and is required to initiate the formation of immature vessels, with this aim, present study was executed to evaluate VEGF expression in kertocystic odontogenic tumor, dentigerous cyst and radicular cyst (RC). A retrospective study was carried out comprising a total of 31 cases; 13 cases of keratocystic odontogenic tumor (KCOT), nine cases of dentigerous cyst (DC) and nine cases of RC. The sections were stained immunohistochemically with VEGF antibody and were evaluated for the presence and intensity of the immuno reactive cells. Statistical analysis was carried out using Chi-square test to inter-compare the VEGF expression between KCOT, DC, and RC. VEGF expression in the epithelium and connective tissue was significantly higher in KCOT compared to dentigerous and RC. One case of KCOT with carcinomatous change also revealed positive results for the VEGF expression in the dysplastic epithelium, tumor islands, and connective tissue. The significant difference was observed on inter-comparison of the VEGF expression in the connective tissue of KCOT and DC, whereas no significant difference was observed in the VEGF expression in the connective tissue of KCOT and DC. The present study data supports the literature finding that angiogenesis can be important in the progression and enlargement of odontogenic cysts similarly to what occurs in neoplastic conditions and further it can be concluded that the higher positivity for VEGF of KCOT could help to

Solid-pseudopapillary neoplasm of the pancreas: Clinicopathologic and immunohistochemical analysis of nine cases

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Banu Yilmaz Ozguven

2015-01-01

Full Text Available Background: Solid-pseudopapillary neoplasm (SPPN of the pancreas is a distinctive tumor of low malignant potential with a predilection for female patients in the second and third decades of life. We studied nine cases of SPPN of the pancreas and reviewed the literature concerning these uncommon tumors. Materials and Methods: A total of 7 cases of SPPN located in the tail of the pancreas and two located in the head of the pancreas were presented. Distal pancreatectomy in three patients and distal pancreatectomy with splenectomy in two patients Whipple′s operation in four patients were performed. Histological diagnosis was made by performing hematoxylin-eosin and periodic acid-Schiff staining, immunohistochemical staining. Follow-up of the patients was between 2 months and 12 years. Results: Computed tomography and magnetic resonance imaging were found as equivocal for diagnosis. Mass containing cystic and solid areas were not characteristic but raised suspicion of SPPN. Pathologic examination showed SPPN in all patients. No metastasis or recurrence was detected during follow-up. Conclusions: Solid-pseudopapillary neoplasm is a relatively rare tumor, and patients tend to survive for a long period. Preoperative imaging is not characteristic. Pathologic examination is the mainstay in the diagnosis. Complete surgical removal is the best choice of treatment.

Validation of tissue microarray technology in squamous cell carcinoma of the esophagus

NARCIS (Netherlands)

Boone, Judith; van Hillegersberg, Richard; van Diest, Paul J.; Offerhaus, G. Johan A.; Borel Rinkes, Inne H. M.; ten Kate, Fiebo J. W.

2008-01-01

Tissue microarray (TMA) technology has been developed to facilitate high-throughput immunohistochemical and in situ hybridization analysis of tissues by inserting small tissue biopsy cores into a single paraffin block. Several studies have revealed novel prognostic biomarkers in esophageal squamous

Biomarker expression in rectal cancer tissue before and after neoadjuvant therapy

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Boogerd LS

2018-03-01

Full Text Available Leonora SF Boogerd,1 Maxime JM van der Valk,1 Martin C Boonstra,1 Hendrica AJM Prevoo,1 Denise E Hilling,1 Cornelis JH van de Velde,1 Cornelis FM Sier,1 Arantza Fariña Sarasqueta,2 Alexander L Vahrmeijer1 1Department of Surgery, 2Department of Pathology, Leiden University Medical Center, Leiden, the Netherlands Purpose: Intraoperative identification of rectal cancer (RC can be challenging, especially because of fibrosis after treatment with preoperative chemo- and radiotherapy (CRT. Tumor-targeted fluorescence imaging can enhance the contrast between tumor and normal tissue during surgery. Promising targets for RC imaging are carcinoembryonic antigen (CEA, epithelial cell adhesion molecule (EpCAM and the tyrosine-kinase receptor Met (c-Met. The effect of CRT on their expression determines their applicability for imaging. Therefore, we investigated whether CRT modifies expression patterns in tumors, lymph node (LN metastases and adjacent normal rectal tissues. Patients and methods: Preoperative biopsies, primary tumor specimens and metastatic LNs were collected from 38 RC patients who did not receive CRT (cohort 1 and 34 patients who did (cohort 2. CEA, EpCAM and c-Met expression was determined using immunohistochemical staining and was semiquantified by a total immunostaining score (TIS, consisting of the percentage and intensity of stained tumor cells (0–12. Results: In both cohorts CEA, EpCAM and c-Met were significantly highly expressed in >60% of tumor tissues compared with adjacent normal epithelium (T/N ratio, P<0.01. EpCAM showed the most homogenous expression in tumors, whereas CEA showed the highest T/N ratio. Most importantly, CEA and EpCAM expression did not significantly change in normal or neoplastic RC tissue after CRT, whereas levels of c-Met changed (P=0.02. Tissues of eight patients with a pathological complete response after CRT showed expression of all biomarkers with TIS close to normal epithelium. Conclusion: Histological

Smoking attenuates wound inflammation and proliferation while smoking cessation restores inflammation but not proliferation

DEFF Research Database (Denmark)

Sørensen, Lars Tue; Toft, Birgitte; Rygaard, Jørgen

2010-01-01

of wounding and excision was repeated after 4, 8, and 12 weeks. All excised tissue was stained with hematoxylin-eosin and immunohistochemically for macrophages (CD68), procollagen 1 N-terminal propeptide (PINP) in fibroblasts, and endothelial cells (CD31). The cellularity was assessed and scored by two...

Image analysis as an adjunct to manual HER-2 immunohistochemical review: a diagnostic tool to standardize interpretation.

LENUS (Irish Health Repository)

Dobson, Lynne

2010-07-01

AIMS: Accurate determination of HER-2 status is critical to identify patients for whom trastuzumab treatment will be of benefit. Although the recommended primary method of evaluation is immunohistochemistry, numerous reports of variability in interpretation have raised uncertainty about the reliability of results. Recent guidelines have suggested that image analysis could be an effective tool for achieving consistent interpretation, and this study aimed to assess whether this technology has potential as a diagnostic support tool. METHODS AND RESULTS: Across a cohort of 275 cases, image analysis could accurately classify HER-2 status, with 91% agreement between computer-aided classification and the pathology review. Assessment of the continuity of membranous immunoreactivity in addition to intensity of reactivity was critical to distinguish between negative and equivocal cases and enabled image analysis to report a lower referral rate of cases for confirmatory fluorescence in situ hybridization (FISH) testing. An excellent concordance rate of 95% was observed between FISH and the automated review across 136 informative cases. CONCLUSIONS: This study has validated that image analysis can robustly and accurately evaluate HER-2 status in immunohistochemically stained tissue. Based on these findings, image analysis has great potential as a diagnostic support tool for pathologists and biomedical scientists, and may significantly improve the standardization of HER-2 testing by providing a quantitative reference method for interpretation.

Conjugated linoleic acid induces apoptosis through estrogen receptor alpha in human breast tissue

International Nuclear Information System (INIS)

Wang, Li-Shu; Huang, Yi-Wen; Liu, Suling; Yan, Pearlly; Lin, Young C

2008-01-01

Conjugated linoleic acid (CLA), a naturally occurring fatty acid found in ruminant products such as milk and beef, has been shown to possess anti-cancer activities in in vivo animal models and in vitro cell culture systems. In human breast cancer, the overall duration of estrogen exposure is the most important risk factor for developing estrogen-responsive breast cancer. Accordingly, it has been suggested that estrogen exposure reduces apoptosis through the up-regulation of the anti-apoptosis protein, Bcl-2. Bcl-2, an anti-apoptotic protein, regulates apoptosis and plays a crucial role in the development and growth regulation of normal and cancerous cells. Our research interest is to examine the effects of CLA on the induction of apoptosis in human breast tissues. The localization of Bcl-2 in both normal and cancerous human breast tissues was determined by immunohistochemical staining and the Bcl-2 protein expression was tested by western blot analysis. Co-culture of epithelial cells and stromal cells was carried out in the presence or absence of CLA to evaluate apoptosis in the context of a cell-cell interaction. The results showed that both normal and cancerous breast tissues were positive for Bcl-2 staining, which was higher overall in mammary ducts but very low in the surrounding stromal compartment. Interestingly, by quantifying the western blot data, basal Bcl-2 protein levels were higher in normal breast epithelial cells than in cancerous epithelial cells. Furthermore, treatment with 17β-estradiol (E 2 ) stimulated growth and up-regulated Bcl-2 expression in estrogen responsive breast epithelial cells; however, these carcinogenic effects were diminished by either CLA or 4-Hydroxytamoxifen (Tam) and were suppressed further by the combination of CLA and Tam. In both one cell type cultured and co-culture systems, CLA induced cell apoptosis in ERα transfected MDA-MB-231 cells but not in the wild type MDA-MB-231 cells. These data, therefore, demonstrate that

Conjugated linoleic acid induces apoptosis through estrogen receptor alpha in human breast tissue

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Liu Suling

2008-07-01

Full Text Available Abstract Background Conjugated linoleic acid (CLA, a naturally occurring fatty acid found in ruminant products such as milk and beef, has been shown to possess anti-cancer activities in in vivo animal models and in vitro cell culture systems. In human breast cancer, the overall duration of estrogen exposure is the most important risk factor for developing estrogen-responsive breast cancer. Accordingly, it has been suggested that estrogen exposure reduces apoptosis through the up-regulation of the anti-apoptosis protein, Bcl-2. Bcl-2, an anti-apoptotic protein, regulates apoptosis and plays a crucial role in the development and growth regulation of normal and cancerous cells. Our research interest is to examine the effects of CLA on the induction of apoptosis in human breast tissues. Methods The localization of Bcl-2 in both normal and cancerous human breast tissues was determined by immunohistochemical staining and the Bcl-2 protein expression was tested by western blot analysis. Co-culture of epithelial cells and stromal cells was carried out in the presence or absence of CLA to evaluate apoptosis in the context of a cell-cell interaction. Results The results showed that both normal and cancerous breast tissues were positive for Bcl-2 staining, which was higher overall in mammary ducts but very low in the surrounding stromal compartment. Interestingly, by quantifying the western blot data, basal Bcl-2 protein levels were higher in normal breast epithelial cells than in cancerous epithelial cells. Furthermore, treatment with 17β-estradiol (E2 stimulated growth and up-regulated Bcl-2 expression in estrogen responsive breast epithelial cells; however, these carcinogenic effects were diminished by either CLA or 4-Hydroxytamoxifen (Tam and were suppressed further by the combination of CLA and Tam. In both one cell type cultured and co-culture systems, CLA induced cell apoptosis in ERα transfected MDA-MB-231 cells but not in the wild type MDA

A rapid method combining Golgi and Nissl staining to study neuronal morphology and cytoarchitecture.

Science.gov (United States)

Pilati, Nadia; Barker, Matthew; Panteleimonitis, Sofoklis; Donga, Revers; Hamann, Martine

2008-06-01

The Golgi silver impregnation technique gives detailed information on neuronal morphology of the few neurons it labels, whereas the majority remain unstained. In contrast, the Nissl staining technique allows for consistent labeling of the whole neuronal population but gives very limited information on neuronal morphology. Most studies characterizing neuronal cell types in the context of their distribution within the tissue slice tend to use the Golgi silver impregnation technique for neuronal morphology followed by deimpregnation as a prerequisite for showing that neuron's histological location by subsequent Nissl staining. Here, we describe a rapid method combining Golgi silver impregnation with cresyl violet staining that provides a useful and simple approach to combining cellular morphology with cytoarchitecture without the need for deimpregnating the tissue. Our method allowed us to identify neurons of the facial nucleus and the supratrigeminal nucleus, as well as assessing cellular distribution within layers of the dorsal cochlear nucleus. With this method, we also have been able to directly compare morphological characteristics of neuronal somata at the dorsal cochlear nucleus when labeled with cresyl violet with those obtained with the Golgi method, and we found that cresyl violet-labeled cell bodies appear smaller at high cellular densities. Our observation suggests that cresyl violet staining is inadequate to quantify differences in soma sizes.

Metallothionein - immunohistochemical cancer biomarker: a meta-analysis.

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Jaromir Gumulec

Full Text Available Metallothionein (MT has been extensively investigated as a molecular marker of various types of cancer. In spite of the fact that numerous reviews have been published in this field, no meta-analytical approach has been performed. Therefore, results of to-date immunohistochemistry-based studies were summarized using meta-analysis in this review. Web of science, PubMed, Embase and CENTRAL databases were searched (up to April 30, 2013 and the eligibility of individual studies and heterogeneity among the studies was assessed. Random and fixed effects model meta-analysis was employed depending on the heterogeneity, and publication bias was evaluated using funnel plots and Egger's tests. A total of 77 studies were included with 8,015 tissue samples (4,631 cases and 3,384 controls. A significantly positive association between MT staining and tumors (vs. healthy tissues was observed in head and neck (odds ratio, OR 9.95; 95% CI 5.82-17.03 and ovarian tumors (OR 7.83; 1.09-56.29, and a negative association was ascertained in liver tumors (OR 0.10; 0.03-0.30. No significant associations were identified in breast, colorectal, prostate, thyroid, stomach, bladder, kidney, gallbladder, and uterine cancers and in melanoma. While no associations were identified between MT and tumor staging, a positive association was identified with the tumor grade (OR 1.58; 1.08-2.30. In particular, strong associations were observed in breast, ovarian, uterine and prostate cancers. Borderline significant association of metastatic status and MT staining were determined (OR 1.59; 1.03-2.46, particularly in esophageal cancer. Additionally, a significant association between the patient prognosis and MT staining was also demonstrated (hazard ratio 2.04; 1.47-2.81. However, a high degree of inconsistence was observed in several tumor types, including colorectal, kidney and prostate cancer. Despite the ambiguity in some tumor types, conclusive results are provided in the tumors of

An empirical analysis of the precision of estimating the numbers of neurons and glia in human neocortex using a fractionator-design with sub-sampling

DEFF Research Database (Denmark)

Lyck, L.; Santamaria, I.D.; Pakkenberg, B.

2009-01-01

Improving histomorphometric analysis of the human neocortex by combining stereological cell counting with immunchistochemical visualisation of specific neuronal and glial cell populations is a methodological challenge. To enable standardized immunohistochemical staining, the amount of brain tissue...... at each level of sampling was determined empirically. The methodology was tested in three brains analysing the contribution of the multi-step sampling procedure to the precision on the estimated total numbers of immunohistochemically defined NeuN expressing (NeuN(+)) neurons and CD45(+) microglia...

The immune cell composition in Barrett's metaplastic tissue resembles that in normal duodenal tissue.

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Alexandra Lind

Full Text Available BACKGROUND AND OBJECTIVE: Barrett's esophagus (BE is characterized by the transition of squamous epithelium into columnar epithelium with intestinal metaplasia. The increased number and types of immune cells in BE have been indicated to be due to a Th2-type inflammatory process. We tested the alternative hypothesis that the abundance of T-cells in BE is caused by a homing mechanism that is found in the duodenum. PATIENTS AND METHODS: Biopsies from BE and duodenal tissue from 30 BE patients and duodenal tissue from 18 controls were characterized by immmunohistochemistry for the presence of T-cells and eosinophils(eos. Ex vivo expanded T-cells were further phenotyped by multicolor analysis using flowcytometry. RESULTS: The high percentage of CD4(+-T cells (69±3% (mean±SEM/n = 17, by flowcytometry, measured by flowcytometry and immunohistochemistry, and the presence of non-activated eosinophils found in BE by immunohistochemical staining, were not different from that found in duodenal tissue. Expanded lymphocytes from these tissues had a similar phenotype, characterized by a comparable but low percentage of αE(CD103 positive CD4(+cells (44±5% in BE, 43±4% in duodenum of BE and 34±7% in duodenum of controls and a similar percentage of granzyme-B(+CD8(+ cells(44±5% in BE, 33±6% in duodenum of BE and 36±7% in duodenum of controls. In addition, a similar percentage of α4β7(+ T-lymphocytes (63±5% in BE, 58±5% in duodenum of BE and 62±8% in duodenum of controls was found. Finally, mRNA expression of the ligand for α4β7, MAdCAM-1, was also similar in BE and duodenal tissue. No evidence for a Th2-response was found as almost no IL-4(+-T-cells were seen. CONCLUSION: The immune cell composition (lymphocytes and eosinophils and expression of intestinal adhesion molecule MAdCAM-1 is similar in BE and duodenum. This supports the hypothesis that homing of lymphocytes to BE tissue is mainly caused by intestinal homing signals rather than to an

Quantification of histochemical stains using whole slide imaging: development of a method and demonstration of its usefulness in laboratory quality control.

Science.gov (United States)

Gray, Allan; Wright, Alex; Jackson, Pete; Hale, Mike; Treanor, Darren

2015-03-01

Histochemical staining of tissue is a fundamental technique in tissue diagnosis and research, but it suffers from significant variability. Efforts to address this include laboratory quality controls and quality assurance schemes, but these rely on subjective interpretation of stain quality, are laborious and have low reproducibility. We aimed (1) to develop a method for histochemical stain quantification using whole slide imaging and image analysis and (2) to demonstrate its usefulness in measuring staining variation. A method to quantify the individual stain components of histochemical stains on virtual slides was developed. It was evaluated for repeatability and reproducibility, then applied to control sections of an appendix to quantify H&E staining (H/E intensities and H:E ratio) between automated staining machines and to measure differences between six regional diagnostic laboratories. The method was validated with laboratories from 0.57 to 0.89. A simple method using whole slide imaging can be used to quantify and compare histochemical staining. This method could be deployed in routine quality assurance and quality control. Work is needed on whole slide imaging devices to improve reproducibility. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Granulocyte-Colony Stimulating Factor Receptor, Tissue Factor, and VEGF-R Bound VEGF in Human Breast Cancer In Loco.

Science.gov (United States)

Wojtukiewicz, Marek Z; Sierko, Ewa; Skalij, Piotr; Kamińska, Magda; Zimnoch, Lech; Brekken, Ralf A; Thorpe, Philip E

2016-01-01

Doxorubicin and docetaxel-based chemotherapy regimens used in breast cancer patients are associated with high risk of febrile neutropenia (FN). Granulocyte colony-stimulating factors (G-CSF) are recommended for both treating and preventing chemotherapy-induced neutropenia. Increased thrombosis incidence in G-CSF treated patients was reported; however, the underlying mechanisms remain unclear. The principal activator of blood coagulation in cancer is tissue factor (TF). It additionally contributes to cancer progression and stimulates angiogenesis. The main proangiogenic factor is vascular endothelial growth factor (VEGF). The aim of the study was to evaluate granulocyte-colony stimulating factor receptor (G-CSFR), tissue factor (TF) expression and vascular endothelial growth factor receptor (VEGF-R) bound VEGF in human breast cancer in loco. G-CSFR, TF and VEGFR bound VEGF (VEGF: VEGFR) were assessed in 28 breast cancer tissue samples. Immunohistochemical (IHC) methodologies according to ABC technique and double staining IHC procedure were employed utilizing antibodies against G-CSFR, TF and VEGF associated with VEGFR (VEGF: VEGFR). Expression of G-CSFR was demonstrated in 20 breast cancer tissue specimens (71%). In 6 cases (21%) the expression was strong (IRS 9-12). Strong expression of TF was observed in all investigated cases (100%). Moreover, expression of VEGF: VEGFR was visualized in cancer cells (IRS 5-8). No presence of G-CSFR, TF or VEGF: VEGFR was detected on healthy breast cells. Double staining IHC studies revealed co-localization of G-CSFR and TF, G-CSFR and VEGF: VEGFR, as well as TF and VEGF: VEGFR on breast cancer cells and ECs. The results of the study indicate that GCSFR, TF and VEGF: VEGFR expression as well as their co-expression might influence breast cancer biology, and may increase thromboembolic adverse events incidence.

Assessment of a novel VEGF targeted agent using patient-derived tumor tissue xenograft models of colon carcinoma with lymphatic and hepatic metastases.

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Ketao Jin

Full Text Available The lack of appropriate tumor models of primary tumors and corresponding metastases that can reliably predict for response to anticancer agents remains a major deficiency in the clinical practice of cancer therapy. It was the aim of our study to establish patient-derived tumor tissue (PDTT xenograft models of colon carcinoma with lymphatic and hepatic metastases useful for testing of novel molecularly targeted agents. PDTT of primary colon carcinoma, lymphatic and hepatic metastases were used to create xenograft models. Hematoxylin and eosin staining, immunohistochemical staining, genome-wide gene expression analysis, pyrosequencing, qRT-PCR, and western blotting were used to determine the biological stability of the xenografts during serial transplantation compared with the original tumor tissues. Early passages of the PDTT xenograft models of primary colon carcinoma, lymphatic and hepatic metastases revealed a high degree of similarity with the original clinical tumor samples with regard to histology, immunohistochemistry, genes expression, and mutation status as well as mRNA expression. After we have ascertained that these xenografts models retained similar histopathological features and molecular signatures as the original tumors, drug sensitivities of the xenografts to a novel VEGF targeted agent, FP3 was evaluated. In this study, PDTT xenograft models of colon carcinoma with lymphatic and hepatic metastasis have been successfully established. They provide appropriate models for testing of novel molecularly targeted agents.

Evaluation of biosafe alternatives as xylene substitutes in hematoxylin and eosin staining procedure: A comparative pilot study

Science.gov (United States)

Sravya, Taneeru; Rao, Guttikonda Venkateswara; Kumari, Masabattula Geetha; Sagar, Yerraguntla Vidya; Sivaranjani, Yeluri; Sudheerkanth, Kondamarri

2018-01-01

Background: Xylene is synthetic hydrocarbon produced from coal tar known for its wide usage as universal solvent which has many hazardous effects. The aim of this study is to compare the efficacy of xylene-free hematoxylin and eosin (H and E) sections with conventional H and E sections. Materials and Methods: The study included ninety paraffin-embedded tissue blocks. Of these, sixty blocks were processed with sesame oil (xylene alternative) and thirty blocks with xylene. The study sample was divided into three groups. Sixty sections which are taken from sesame oil-processed blocks were stained with xylene-free H and E staining method. In xylene-free staining method, 95% diluted lemon water (Group A) and 1.7% dish washing solution (DWS, Group B) were used as deparaffinizing agents whereas the remaining 30 sections were processed with xylene and stained with conventional H and E staining method (Group C). Slides were scored for the following parameters: (i) nuclear staining (adequate = score 1, inadequate = score 0), (ii) cytoplasmic staining (adequate = score 1, inadequate = score 0), (iii) uniformity (present = score 1, absent = score 0), (iv) clarity (present = score 1, absent = score 0) and (v) intensity (present = score 1, absent = score 0). Score ≤2 was considered inadequate for diagnosis while scores 3–5 were considered adequate for diagnosis. Results: Adequate nuclear staining was noted in 90% of sections of Group A and 100% each in Group B and Group C (P 0.05); adequate uniformity of staining in 53.3% of sections of Group A, 70% in Group B and 83.3% in Group C (P 0.05) and adequate intensity of staining in 76.7% sections of Group A, 93.3% in Group B and 100% in Group C (P < 0.05). Group C sections stained adequate for diagnosis (93.3%) followed by Group B (88.7%) and Group A (78%; P < 0.05). Conclusion: Tissues processed with sesame oil and stained using 1.7% DWS were found to be effective alternative to xylene.

Cell wall staining with Trypan blue enables quantitative analysis of morphological changes in yeast cells

DEFF Research Database (Denmark)

Liesche, Johannes; Marek, Magdalena; Günther-Pomorski, Thomas

2015-01-01

staining with fluorescent dyes is a valuable tool. Furthermore, cell wall staining is used to facilitate sub-cellular localization experiments with fluorescently-labeled proteins and the detection of yeast cells in non-fungal host tissues. Here, we report staining of Saccharomyces cerevisiae cell wall......Yeast cells are protected by a cell wall that plays an important role in the exchange of substances with the environment. The cell wall structure is dynamic and can adapt to different physiological states or environmental conditions. For the investigation of morphological changes, selective...... with Trypan Blue, which emits strong red fluorescence upon binding to chitin and yeast glucan; thereby, it facilitates cell wall analysis by confocal and super-resolution microscopy. The staining pattern of Trypan Blue was similar to that of the widely used UV-excitable, blue fluorescent cell wall stain...

Partial characterization of insulin-like growth factor I in primary human lung cancers using immunohistochemical and receptor autoradiographic techniques

International Nuclear Information System (INIS)

Shigematsu, K.; Kataoka, Y.; Kamio, T.; Kurihara, M.; Niwa, M.; Tsuchiyama, H.

1990-01-01

We investigated primary human lung cancers resected surgically or obtained at autopsy. Included were squamous cell carcinoma (SQC) (five cases), adenocarcinoma (ADC) (six cases), large cell carcinoma (LCC) (four cases), and small cell carcinoma (SCC) (two cases). The objective of the study was to search for the presence of insulin-like growth factor I (IGF-I)-like immunoreactivity using immunohistochemical staining and for the localization of IGF-I binding sites, using in vitro quantitative receptor autoradiographic techniques. IGF-I-like immunostaining was present in all cases of SQC, ADC, and LCC, but not in cases of SCC. Strong immunostaining was observed in cases of SQC. On the other hand, ADC and LCC tissues showed a moderate or weak staining. Specific binding sites for IGF-I were present in all cases of SQC, ADC, LCC, and SCC examined. High densities of 125I-IGF-I binding sites were localized in cases of SQC and SCC. Low to high densities of the binding sites were found in LCC. Cases of ADC showed low densities of 125I-IGF-I binding sites. Specific binding obtained at a concentration of 80 pM 125I-IGF-I was competitively displaced by unlabeled IGF-I, with a 50% inhibitory concentration value of 1.84 +/- 0.31 x 10(-10) mol, whereas human insulin was much less potent in displacing the binding. This specificity profile is consistent with characteristics of IGF-I receptors. Scatchard analysis showed the presence of a single class of high affinity binding sites for IGF-I, with a Kd of approximately 1 nmol. Thus, the possibility that IGF-I may play a role in the growth of human lung cancers would have to be considered

Stain Deconvolution Using Statistical Analysis of Multi-Resolution Stain Colour Representation.

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Najah Alsubaie

Full Text Available Stain colour estimation is a prominent factor of the analysis pipeline in most of histology image processing algorithms. Providing a reliable and efficient stain colour deconvolution approach is fundamental for robust algorithm. In this paper, we propose a novel method for stain colour deconvolution of histology images. This approach statistically analyses the multi-resolutional representation of the image to separate the independent observations out of the correlated ones. We then estimate the stain mixing matrix using filtered uncorrelated data. We conducted an extensive set of experiments to compare the proposed method to the recent state of the art methods and demonstrate the robustness of this approach using three different datasets of scanned slides, prepared in different labs using different scanners.

Double-tracer autoradiography with Cu-ATSM/FDG and immunohistochemical interpretation in four different mouse implanted tumor models

Energy Technology Data Exchange (ETDEWEB)

Tanaka, Takeshi [Department of Otorhinolaryngology, University of Fukui, Matsuoka, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan)]. E-mail: wplants@mac.com; Furukawa, Takako [Biomedical Imaging Research Center, University of Fukui, Matsuoka, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan); Fujieda, Shigeharu [Department of Otorhinolaryngology, University of Fukui, Matsuoka, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan); Kasamatsu, Shingo [Biomedical Imaging Research Center, University of Fukui, Matsuoka, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan); Yonekura, Yoshiharu [Biomedical Imaging Research Center, University of Fukui, Matsuoka, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan); Fujibayashi, Yasuhisa [Biomedical Imaging Research Center, University of Fukui, Matsuoka, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan)]. E-mail: yfuji@fmsrsa.fukui-med.ac.jp

2006-08-15

Background: We studied the regional characteristics within tumor masses using PET tracers and immunohistochemical methods. Methods: The intratumoral distribution of {sup 64}Cu-diacetyl-bis(N4-methylthiosemicarbazone) ([{sup 64}Cu]Cu-ATSM) and [{sup 18}F] 2-fluoro-2-deoxyglucose ({sup 18}F]FDG) in mice with tumors of four different origins (LLC1, Meth-A, B16 and colon26) was compared with the immunohistochemical staining of proliferating cells (Ki67), blood vessels (CD34 or von Willebrand factor), and apoptotic cells (terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling method). Results: With all cell lines, [{sup 64}Cu]Cu-ATSM and [{sup 18}F]FDG were distributed with different gradation in the tumor mass. The immunohistochemical study demonstrated that the high [{sup 64}Cu]Cu-ATSM uptake regions were hypovascular and consisted of tumor cells arrested in the cell cycle, whereas the high [{sup 18}F]FDG uptake regions were hypervascular and consisted of proliferating cells. Conclusion: In our study, it was revealed that one tumor mass contained two regions with different characteristics, which could be distinguished by [{sup 64}Cu]Cu-ATSM and [{sup 18}F]FDG. Because hypoxia and cell cycle arrest are critical factors to reduce tumor sensitivity to radiation and conventional chemotherapy, regions with such characteristics should be treated intensively as one of the primary targets. [{sup 64}Cu]Cu-ATSM, which can delineate hypoxic and cell cycle-arrested regions in tumors, may provide valuable information for cancer treatment as well as possibly for treating such regions directly as an internal radiotherapy reagent.

Radiation induced cell death in cervical squamous cell carcinoma. An immunohistochemical and ultrastructural study

International Nuclear Information System (INIS)

Atari, Eio; Toda, Takayoshi; Sadi, A.M.; Egawa, Haruhiko; Moromizato, Hidehiko; Mamadi, T.; Kiyuna, Masaya

1998-01-01

To study the process of cell death in cervical squamous cell carcinoma (SCC) after radiation, an ultrastructural and immunohistochemical study was performed. Paraffin-embedded tissue blocks of biopsy samples pre- and post-radiation stage III SCC (n=15) were collected. Irradiation caused varying ultrastructural changes including nuclear and cytoplasmic disorganization suggesting cell necrosis. Immunohistochemically, the pre-radiation specimens showed no positive reaction for tumor necrosis factor-alpha (TNF-α), tumor necrosis factor-receptor (TNF-γ) or Fas. C-fos, p53 and bcl-2 showed positive reactions in only a few non-irradiated specimens. All of the irradiated specimens showed a positive reaction for TNF-α, and variable positive reactions were observed for TNF-γ, Fas, p53, c-fos and bcl-2. These results suggest that TNF-α, TNF-γ, and c-fos are responsible for radiation induced cell death in cervical SCC. (author)

Finding an optimum immuno-histochemical feature set to distinguish benign phyllodes from fibroadenoma.

Science.gov (United States)

Maity, Priti Prasanna; Chatterjee, Subhamoy; Das, Raunak Kumar; Mukhopadhyay, Subhalaxmi; Maity, Ashok; Maulik, Dhrubajyoti; Ray, Ajoy Kumar; Dhara, Santanu; Chatterjee, Jyotirmoy

2013-05-01

Benign phyllodes and fibroadenoma are two well-known breast tumors with remarkable diagnostic ambiguity. The present study is aimed at determining an optimum set of immuno-histochemical features to distinguish them by analyzing important observations on expressions of important genes in fibro-glandular tissue. Immuno-histochemically, the expressions of p63 and α-SMA in myoepithelial cells and collagen I, III and CD105 in stroma of tumors and their normal counterpart were studied. Semi-quantified features were analyzed primarily by ANOVA and ranked through F-scores for understanding relative importance of group of features in discriminating three classes followed by reduction in F-score arranged feature space dimension and application of inter-class Bhattacharyya distances to distinguish tumors with an optimum set of features. Among thirteen studied features except one all differed significantly in three study classes. F-Ranking of features revealed highest discriminative potential of collagen III (initial region). F-Score arranged feature space dimension and application of Bhattacharyya distance gave rise to a feature set of lower dimension which can discriminate benign phyllodes and fibroadenoma effectively. The work definitely separated normal breast, fibroadenoma and benign phyllodes, through an optimal set of immuno-histochemical features which are not only useful to address diagnostic ambiguity of the tumors but also to spell about malignant potentiality. Copyright © 2013 Elsevier Ltd. All rights reserved.

Immunohistochemical detection of interleukin-8 in inflamed porcine tissues

DEFF Research Database (Denmark)

Laursen, Henriette; Jensen, Henrik Elkær; Leifsson, Páll S.

2014-01-01

The objective of this study was to identify the specific localization of interleukin-8 (IL-8) in cells in situ in a variety of inflammatory processes in different tissues from pigs. Our hypothesis was that IL-8 primarily is a neutrophil related cytokine present in all extravascular neutrophils...... while expression also occurs in other cell types in response to an inflammatory stimulus. Using IL-8 immunohistochemistry we discovered that neutrophils were the predominant IL-8 positive cell population while epithelial cell types and endothelium of postcapillary venules could be positive when situated...... in close vicinity of an inflammatory lesion. Furthermore, endothelial cells of newly formed vessels in granulation tissue were positive in some specimens. Some sub-populations of inflammatory neutrophils were, however, IL-8 negative which could reflect some phase of neutrophil swarming....

Learning Histopathological Patterns

DEFF Research Database (Denmark)

Kårsnäs, Andreas; Dahl, Anders Lindbjerg; Larsen, Rasmus

2011-01-01

trained from representative image samples. Nuclei counting is based on a nucleus model that takes size, shape and nucleus probability into account. We obtain the probability of a nucleus from our segmentation procedure. Our method is experimentally validated on images stained with nuclear markers......We propose a technique for analyzing images of immunohistochemically stained tissue samples for extracting features that correlate with patient disease. We address the problem of quantifying tumor tissue and segmenting and counting nuclei. Our method utilizes a flexible segmentation technique...... for the Estrogen Receptor (ER) and proliferation marker KI-67. In addition we qualitatively validate our method for tumor tissue segmentation and we obtain state of the art results on cell nuclei separation....

[Immunocytochemical demonstration of astrocytes in brain sections combined with Nissl staining].

Science.gov (United States)

Korzhevskiĭ, D E; Otellin, V A

2004-01-01

The aim of the present study was to develop an easy and reliable protocol of combined preparation staining, which would unite the advantages of immunocytochemical demonstration of astrocytes with the availability to evaluate functional state of neurons provided by Nissl technique. The presented protocol of paraffin sections processing allows to retain high quality of tissue structure and provides for selective demonstration of astrocytes using the monoclonal antibodies against glial fibrillary acidic protein and contrast Nissl staining of cells. The protocol can be used without any changes for processing of brain sections obtained from the humans and other mammals with the exception of mice and rabbits.

Effect of Brain Tumor Presence During Radiation on Tissue Toxicity: Transcriptomic and Metabolic Changes.

Science.gov (United States)

Zawaski, Janice A; Sabek, Omaima M; Voicu, Horatiu; Eastwood Leung, Hon-Chiu; Gaber, M Waleed

2017-11-15

Radiation therapy (RT) causes functional and transcriptomic changes in the brain; however, most studies have been carried out in normal rodent brains. Here, the long-term effect of irradiation and tumor presence during radiation was investigated. Male Wistar rats ∼7 weeks old were divided into 3 groups: sham implant, RT+sham implant, and RT+tumor implant (C6 glioma). Hypofractionated irradiation (8 or 6 Gy/day for 5 days) was localized to a 1-cm strip of cranium starting 5 days after implantation, resulting in complete tumor regression and prolonged survival. Biopsy of tissue was performed in the implant area 65 days after implantation. RNA was hybridized to GeneChip Rat Exon 1.0 ST array. Data were analyzed using significant analysis of microarrays and ingenuity pathway analysis. 1 H magnetic resonance spectroscopy ( 1 H-MRS) imaging was performed in the implantation site 65 to 70 days after implantation using a 9.4 T Biospec magnetic resonance imaging scanner with a quadrature rat brain array. Immunohistochemical staining for astrogliosis, HMG-CoA synthase 2, γ-aminobutyric acid (GABA) and taurine was performed at ∼65 days after implantation. Eighty-four genes had a false discovery rate <3.5%. We compared RT+tumor implant with RT+sham implant animals. The tumor presence affected networks associated with cancer/cell morphology/tissue morphology. 1 H-MRS showed significant reduction in taurine levels (P<.04) at the implantation site in both groups. However, the RT+tumor group also showed significant increase in levels of neurotransmitter GABA (P=.02). Hippocampal taurine levels were only significantly reduced in the RT+tumor group (P=.03). HMG-CoA synthase 2, GABA and taurine levels were confirmed using staining. Glial fibrillary acidic protein staining demonstrated a significant increase in inflammation that was heightened in the RT+tumor group. Our data indicate that tumor presence during radiation significantly affects long-term functional

Technical Note: Immunohistochemical evaluation of mouse brain irradiation targeting accuracy with 3D-printed immobilization device

Energy Technology Data Exchange (ETDEWEB)

Zarghami, Niloufar, E-mail: nzargham@uwo.ca; Jensen, Michael D. [Department of Medical Biophysics, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); Talluri, Srikanth; Dick, Frederick A. [Department of Biochemistry, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); London Regional Cancer Program, London Health Sciences Centre, 800 Commissioners Road East, London, Ontario N6A 5W9 (Canada); Foster, Paula J. [Imaging Research Laboratories, Robarts Research Institute, 100 Perth Drive, London, Ontario N6A 5K8 (Canada); Department of Medical Biophysics, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); Chambers, Ann F. [Department of Medical Biophysics, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); Department of Oncology, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); London Regional Cancer Program, London Health Sciences Centre, 800 Commissioners Road East, London, Ontario N6A 5W9 (Canada); Wong, Eugene [Department of Physics and Astronomy, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); Department of Medical Biophysics, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); Department of Oncology, The University of Western Ontario, 1151 Richmond Street, London, Ontario N6A 3K7 (Canada); London Regional Cancer Program, London Health Sciences Centre, 800 Commissioners Road East, London, Ontario N6A 5W9 (Canada)

2015-11-15

Purpose: Small animal immobilization devices facilitate positioning of animals for reproducible imaging and accurate focal radiation therapy. In this study, the authors demonstrate the use of three-dimensional (3D) printing technology to fabricate a custom-designed mouse head restraint. The authors evaluate the accuracy of this device for the purpose of mouse brain irradiation. Methods: A mouse head holder was designed for a microCT couch using CAD software and printed in an acrylic based material. Ten mice received half-brain radiation while positioned in the 3D-printed head holder. Animal placement was achieved using on-board image guidance and computerized asymmetric collimators. To evaluate the precision of beam localization for half-brain irradiation, mice were sacrificed approximately 30 min after treatment and brain sections were stained for γ-H2AX, a marker for DNA breaks. The distance and angle of the γ-H2AX radiation beam border to longitudinal fissure were measured on histological samples. Animals were monitored for any possible trauma from the device. Results: Visualization of the radiation beam on ex vivo brain sections with γ-H2AX immunohistochemical staining showed a sharp radiation field within the tissue. Measurements showed a mean irradiation targeting error of 0.14 ± 0.09 mm (standard deviation). Rotation between the beam axis and mouse head was 1.2° ± 1.0° (standard deviation). The immobilization device was easily adjusted to accommodate different sizes of mice. No signs of trauma to the mice were observed from the use of tooth block and ear bars. Conclusions: The authors designed and built a novel 3D-printed mouse head holder with many desired features for accurate and reproducible radiation targeting. The 3D printing technology was found to be practical and economical for producing a small animal imaging and radiation restraint device and allows for customization for study specific needs.

Technical Note: Immunohistochemical evaluation of mouse brain irradiation targeting accuracy with 3D-printed immobilization device

International Nuclear Information System (INIS)

Zarghami, Niloufar; Jensen, Michael D.; Talluri, Srikanth; Dick, Frederick A.; Foster, Paula J.; Chambers, Ann F.; Wong, Eugene

2015-01-01

Purpose: Small animal immobilization devices facilitate positioning of animals for reproducible imaging and accurate focal radiation therapy. In this study, the authors demonstrate the use of three-dimensional (3D) printing technology to fabricate a custom-designed mouse head restraint. The authors evaluate the accuracy of this device for the purpose of mouse brain irradiation. Methods: A mouse head holder was designed for a microCT couch using CAD software and printed in an acrylic based material. Ten mice received half-brain radiation while positioned in the 3D-printed head holder. Animal placement was achieved using on-board image guidance and computerized asymmetric collimators. To evaluate the precision of beam localization for half-brain irradiation, mice were sacrificed approximately 30 min after treatment and brain sections were stained for γ-H2AX, a marker for DNA breaks. The distance and angle of the γ-H2AX radiation beam border to longitudinal fissure were measured on histological samples. Animals were monitored for any possible trauma from the device. Results: Visualization of the radiation beam on ex vivo brain sections with γ-H2AX immunohistochemical staining showed a sharp radiation field within the tissue. Measurements showed a mean irradiation targeting error of 0.14 ± 0.09 mm (standard deviation). Rotation between the beam axis and mouse head was 1.2° ± 1.0° (standard deviation). The immobilization device was easily adjusted to accommodate different sizes of mice. No signs of trauma to the mice were observed from the use of tooth block and ear bars. Conclusions: The authors designed and built a novel 3D-printed mouse head holder with many desired features for accurate and reproducible radiation targeting. The 3D printing technology was found to be practical and economical for producing a small animal imaging and radiation restraint device and allows for customization for study specific needs

The impact of pituicytoma on hormone secretary function

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Ting-Wei Chang

2018-06-01

Full Text Available Background: Pituicytes are glial cells supporting hypothalamic axons in the posterior pituitary gland. Pituicytoma arises from pituicytes, and is currently classified as a World Health Organization grade I central nervous system tumor. Clinically, some patients with pituicytoma are misdiagnosed with functioning pituitary adenoma before surgical interventions and pathological examinations. This study was designed to investigate the correlation between pituicytoma and abnormal hormone function. Methods: The records of patients who underwent pituitary surgery at Linkou Chang Gung Memorial Hospital between 2000 and 2016 were reviewed. Patients with a pathological diagnosis of pituicytoma were included. Among these patients, those with inadequate data and/or specimens were excluded. Clinical information including presenting symptoms, serum hormone level, neuroimages, and results of pathological examinations including hematoxylin and eosin stains and immunohistochemical stains were collected for analysis. Results: Between 2000 and 2016, eight patients were pathologically diagnosed with pituicytoma. Two patients were excluded due to missing data and/or specimens. Among the six remaining patients, five presented with hormone changes. The immunohistochemical stains of pituicytoma confirmed no secretory function; however, the surrounding pituitary glandular tissue showed positive results responsible for clinical hormone changes. Conclusions: Pituicytoma had a negative endocrine secretory function in this study. However, pituicytoma was associated with hyperactivity of the surrounding pituitary glandular tissue, which resulted in clinically abnormal hormone secretion. The diagnosis of pituicytoma before a thorough pathological examination is difficult when the patients present with hormone dysfunction. Therefore, maximum resection of the pituitary tumor is necessary to make the diagnosis of pituicytoma and to exclude pituitary adenoma. Keywords

Combined Staining Techniques for Demonstration of Staphylococcus aureus Biofilm in Routine Histopathology

DEFF Research Database (Denmark)

Jensen, Louise Kruse; Henriksen, Nicole Lind; Bjarnsholt, Thomas

2018-01-01

Aim: Visualization of Staphylococcus aureus biofilm using histochemical staining and combined histochemistry (HC) and immunohistochemistry (IHC). Methods: The ability of S. aureus S54F9 to form biofilm was tested in vitro. Hereafter, infected bone tissue was collected from two different porcine m...

Lectin immunohistochemical evaluation of human bladder carcinomas. A comparison of Carnoy's and formalin fixation.

Science.gov (United States)

Okamura, T; Ueda, K; Ohtaguro, K; Inoue, K; Washida, H; Mori, M; Tatemoto, Y; Fukushima, S

1993-10-01

A lectin immunohistochemical analysis of 51 human bladder carcinomas, including 44 cases of transitional cell carcinoma (TCC) (G1, 15 cases; G2, 17 cases; G3, 12 cases) and 7 cases of squamous cell carcinoma (SCC), was performed. Tissues were obtained by cold punch biopsies, fixed in Carnoy's or 10% formalin solution, stained for binding of 10 different lectins, and evaluated under the light microscope. The lectins used were concanavalin agglutinin (Con A), soybean agglutinin (SBA), Lotus tetragonolobus agglutinin (LTA), Dolichos biflorusa agglutinin (DBA), peanut agglutinin (PNA), Ricinus communis agglutinin I (RCA1), Ulex europaeus agglutinin I, II (UEA-I, II), wheat germ agglutinin (WGA), and Pisum sativum agglutinin (PEA). TCC prepared with Carnoy's fixation tended to show moderately positive Con A, UEA-I, and WGA reactions for G1, and strongly positive reactions for G2 and G3 lesions. UEA-II was mainly negative in G1, but tended to increase to become moderate in G3. DBA tended to show a moderately positive reaction in G1 and G2, but was mainly negative in G3. With formalin fixation, only RCA1 demonstrated grade specific variation, tendency to react moderately in the G1 and G2 cases, and strongly in G3. There were no further differences among the histopathological grades of TCC for other lectins. Thus, Carnoy's fixation appears superior for distinguishing between grades of lesions. SCC tended to react more strongly than TCC with all the various lectins except PEA, independent of fixation.

Richly innervated soft tissues covering the superficial aspect of the extensor origin in patients with chronic painful tennis elbow – Implication for treatment?

Science.gov (United States)

Spang, C.; Alfredson, H.

2017-01-01

Background: Tennis elbow is difficult to treat. The results of surgical treatments are not convincing. Treatment studies on Achilles and patellar tendinopathy targeting the richly innervated and vascularized soft tissues outside the tendon have shown promising outcomes. The innervation patterns in the fibrous/fatty tissues superficially to the elbow extensor origin have not been clarified. Methods: Nine tissue specimens from the fibrous/fatty tissue covering the extensor origin was taken from seven patients (mean age: 45 years) undergoing surgical treatment for chronic painful tennis elbow. The specimens were stained for morphology (haematoxylin & eosin, H&E) and immunohistochemically for general nerve marker protein gene product 9.5 (PGP 9.5) and markers for sympathetic (tyrosine hydroxylase, TH) and sensory nerve fibres (calcitonin gene-related peptide, CGRP). Results: All specimens contained multiple blood vessels and nerve structures indicated by morphology and immunoreactions. There was a frequent occurrence of TH reactions, especially peri-vascularly, but also in nerve fascicles. Immunoreactions for CGRP were seen in nerve fascicles and isolated nerve fibres. Conclusion: The results provide new information on the innervation patterns of the superficial tissues of the extensor origin and their potential as source of tennis elbow pain. Level of Evidence: IV. PMID:28574416

Periostin: a promising target of therapeutical intervention for prostate cancer

Directory of Open Access Journals (Sweden)

Ding Weihong

2011-06-01

Full Text Available Abstract Background In our recent study, Periostin was up-regulated in prostate cancer(PCa compared with benign prostate hyperplasia (BPH by proteomics analysis of prostate biopsies. We investigated the effect of sliencing Periostin by RNA interference (RNAi on the proliferation and migration of PCa LNCap cell line. Methods All the prostate biopsies from PCa, BPH and BPH with local prostatic intraepithelial neoplasm(PIN were analyzed by iTRAQ(Isobaric tags for relative and absolute quantification technology. Western blotting and immunohistochemical staining were used to verify Periostin expression in the tissues of PCa. Periostin expression in different PCa cell lines was determined by immunofluorescence staining, western blotting and reverse transcription PCR(RT-PCR. The LNCap cells with Periostin expression were used for transfecting shRNA-Periostin lentiviral particles. The efficancy of transfecting shRNA lentiviral particles was evaluated by immunofluorescence, western blotting and Real-time PCR. The effect of silencing Periostin expression by RNAi on proliferation of LNCap cells was determined by MTT assay and tumor xenografts. The tissue slices from theses xenografts were analyzed by hematoxylin and eosin(HE staining. The expression of Periostin in the xenografts was deteminned by Immunohistochemical staining and western blotting. The migration of LNCap cells after silencing Periostin gene expression were analyzed in vitro. Results Periostin as the protein of interest was shown 9.12 fold up-regulation in PCa compared with BPH. The overexpression of Periostin in the stroma of PCa was confirmed by western blotting and immunohistochemical staining. Periostin was only expressed in PCa LNCap cell line. Our results indicated that the transfection ratio was more than 90%. As was expected, both the protein level and mRNA level of Periostin in the stably expressing shRNA-Periostin LNCap cells were significantly reduced. The stably expressing sh

A Deep Learning Approach to Digitally Stain Optical Coherence Tomography Images of the Optic Nerve Head.

Science.gov (United States)

Devalla, Sripad Krishna; Chin, Khai Sing; Mari, Jean-Martial; Tun, Tin A; Strouthidis, Nicholas G; Aung, Tin; Thiéry, Alexandre H; Girard, Michaël J A

2018-01-01

To develop a deep learning approach to digitally stain optical coherence tomography (OCT) images of the optic nerve head (ONH). A horizontal B-scan was acquired through the center of the ONH using OCT (Spectralis) for one eye of each of 100 subjects (40 healthy and 60 glaucoma). All images were enhanced using adaptive compensation. A custom deep learning network was then designed and trained with the compensated images to digitally stain (i.e., highlight) six tissue layers of the ONH. The accuracy of our algorithm was assessed (against manual segmentations) using the dice coefficient, sensitivity, specificity, intersection over union (IU), and accuracy. We studied the effect of compensation, number of training images, and performance comparison between glaucoma and healthy subjects. For images it had not yet assessed, our algorithm was able to digitally stain the retinal nerve fiber layer + prelamina, the RPE, all other retinal layers, the choroid, and the peripapillary sclera and lamina cribrosa. For all tissues, the dice coefficient, sensitivity, specificity, IU, and accuracy (mean) were 0.84 ± 0.03, 0.92 ± 0.03, 0.99 ± 0.00, 0.89 ± 0.03, and 0.94 ± 0.02, respectively. Our algorithm performed significantly better when compensated images were used for training (P deep learning algorithm can simultaneously stain the neural and connective tissues of the ONH, offering a framework to automatically measure multiple key structural parameters of the ONH that may be critical to improve glaucoma management.

Imaging Amyloid Tissues Stained with Luminescent Conjugated Oligothiophenes by Hyperspectral Confocal Microscopy and Fluorescence Lifetime Imaging.

Science.gov (United States)

Nyström, Sofie; Bäck, Marcus; Nilsson, K Peter R; Hammarström, Per

2017-10-20

Proteins that deposit as amyloid in tissues throughout the body can be the cause or consequence of a large number of diseases. Among these we find neurodegenerative diseases such as Alzheimer's and Parkinson's disease afflicting primarily the central nervous system, and systemic amyloidosis where serum amyloid A, transthyretin and IgG light chains deposit as amyloid in liver, carpal tunnel, spleen, kidney, heart, and other peripheral tissues. Amyloid has been known and studied for more than a century, often using amyloid specific dyes such as Congo red and Thioflavin T (ThT) or Thioflavin (ThS). In this paper, we present heptamer-formyl thiophene acetic acid (hFTAA) as an example of recently developed complements to these dyes called luminescent conjugated oligothiophenes (LCOs). hFTAA is easy to use and is compatible with co-staining in immunofluorescence or with other cellular markers. Extensive research has proven that hFTAA detects a wider range of disease associated protein aggregates than conventional amyloid dyes. In addition, hFTAA can also be applied for optical assignment of distinct aggregated morphotypes to allow studies of amyloid fibril polymorphism. While the imaging methodology applied is optional, we here demonstrate hyperspectral imaging (HIS), laser scanning confocal microscopy and fluorescence lifetime imaging (FLIM). These examples show some of the imaging techniques where LCOs can be used as tools to gain more detailed knowledge of the formation and structural properties of amyloids. An important limitation to the technique is, as for all conventional optical microscopy techniques, the requirement for microscopic size of aggregates to allow detection. Furthermore, the aggregate should comprise a repetitive β-sheet structure to allow for hFTAA binding. Excessive fixation and/or epitope exposure that modify the aggregate structure or conformation can render poor hFTAA binding and hence pose limitations to accurate imaging.

Methamphetamine and amphetamine concentrations in postmortem rabbit tissues.

Science.gov (United States)

Nagata, T; Kimura, K; Hara, K; Kudo, K

1990-11-01

The feasibility of detecting methamphetamine and its major metabolite, amphetamine, in postmortem tissues over a 2-year period was examined. It is important to determine if the abuse and toxic effects of drugs can be proved from evidence found in decayed, submerged, or stained tissue materials. The blood, urine, liver, skeletal muscle, skin and extremity bones from rabbits given methamphetamine intravenously were kept at room temperature, under 4 different conditions: sealed in a test tube, dried in the open air, submerged in tap water and stained on gauze. Methamphetamine was present in all the samples, with slight change in concentration in case of sealed and air dried tissues. Changes varied in bones kept in water. There were considerable decreases in methamphetamine in blood and urine stains. Despite long term storage, drug abuse and/or toxicity could be determined, in all tissues examined.

Morphological Characterisation of Unstained and Intact Tissue Micro-architecture by X-ray Computed Micro- and Nano-Tomography

Science.gov (United States)

Walton, Lucy A.; Bradley, Robert S.; Withers, Philip J.; Newton, Victoria L.; Watson, Rachel E. B.; Austin, Clare; Sherratt, Michael J.

2015-05-01

Characterisation and quantification of tissue structures is limited by sectioning-induced artefacts and by the difficulties of visualising and segmenting 3D volumes. Here we demonstrate that, even in the absence of X-ray contrast agents, X-ray computed microtomography (microCT) and nanotomography (nanoCT) can circumvent these problems by rapidly resolving compositionally discrete 3D tissue regions (such as the collagen-rich adventitia and elastin-rich lamellae in intact rat arteries) which in turn can be segmented due to their different X-ray opacities and morphologies. We then establish, using X-ray tomograms of both unpressurised and pressurised arteries that intra-luminal pressure not only increases lumen cross-sectional area and straightens medial elastic lamellae but also induces profound remodelling of the adventitial layer. Finally we apply microCT to another human organ (skin) to visualise the cell-rich epidermis and extracellular matrix-rich dermis and to show that conventional histological and immunohistochemical staining protocols are compatible with prior X-ray exposure. As a consequence we suggest that microCT could be combined with optical microscopy to characterise the 3D structure and composition of archival paraffin embedded biological materials and of mechanically stressed dynamic tissues such as the heart, lungs and tendons.

Differential expression of GSK3β and pS9GSK3β in normal human tissues: can pS9GSK3β be an epithelial marker?

Science.gov (United States)

Lee, Hojung; Ro, Jae Y

2015-01-01

Glycogen synthase kinase 3β (GSK3β) and phosphorylated GSK3β at Ser9 (pS9GSK3β) are crucial in cellular proliferation and metabolism. GSK3β and pS9GSK3β are deregulated in many diseases including tumors. Data on altered expression of GSK3β and pS9GSK3β are mainly limited to tumor tissues, thus the expression of GSK3β and pS9GSK3β in normal human tissue has been largely unknown. Thus, we examined the immunohistochemical localization of GSK3β and pS9GSK3β in human fetal and adult tissues, and also compared the expression pattern of GSK3β and pS9GSK3β with that of the CK7 and CK20. We found GSK3β expression in neurons of brain, myenteric plexus in gastrointestinal tract, squamous epithelium of skin, and mammary gland. The expression of pS9GSK3β was restricted to the epithelial cells of breast and pancreaticobiliary duct, distal nephron of kidney, gastrointestinal tract, fallopian tube, epididymis, secretory cell of prostatic gland, and umbrella cell of urinary tract. The staining pattern of pS9GSK3β and CK7 was overlapped in most organs except for gastrointestinal tract where CK7 was negative and CK20 was positive. Our results show that the expression of GSK3β may be associated with differentiation of ectodermal derived tissues and pS9GSK3β with that of epithelial cells of endodermal derived tissues in human. In addition, the expression of pS9GSK3β in the selective epithelial cells may indicate its association with secretory or barrier function of specific cells and may serve as another immunohistochemical marker for epithelial cells.

Pleural and Pulmonary Staining at Inferior Phrenic Arteriography Mimicking a Tumor Staining of Hepatocellular Carcinoma

International Nuclear Information System (INIS)

Lee, Deok Hee; Hwang, Jae Cheol; Lim, Soo Mee; Yoon, Hyun-Ki; Sung, Kyu-Bo; Song, Ho-Young