lesions. Histopathology allows studying the structure and function of cells, tissues, organs and organ systems. stained with HE&S (hematoxylin-eosin and saffron) or can be used for other purposes (special stains, immunohistochemistry, in situ hybridization, etc.). Additionally four of the more difficult tissue/sample collection techniques are demonstrated. Tissue water bath with a thermometer. All test tube contents were well mixed and allowed to stand at 25°C for a further 30 min. 4 0 obj exact angle is generally found by trial and error. and eyes using a modified Davidson’s fluid: Comparison with Bouin’s fluid and conventional, approaches for the evaluation of testicular and, Rousseaux, C.G., and Wallig, M.A., eds), V. Academic Press, San Diego New York Boston. Xylene substitutes are preferred, but if haz-, ards of xylene are well documented, its substitutes have, plastic or resin polymers that should be considered when par-, affin embedding is not appropriate (i.e., methyl methacrylate, to section nondecalcified bone, Epon to obtain very thin sec-, equipment. Abstract:- Onion (Allium cepa L.) is one of the most widely consumed and cultivated vegetable crops in the world, which are effective antioxidants due to their capability to scavenge free radicals. 5% DMSO sterile water; Group IV rats were orally pretreated with 10 ml/kg/day The animals were kept at the animal research facility of the Department of Biomedical Sciences, University of Cape Coast (UCC). Microscopic Evaluation. The genus Allanblackia is a member of the flowering plant family Clusiaceae, which comprises 14 genera and almost 600 species of trees or shrubs that are mainly distributed in tropical regions of the world [10]. Routinely, tissues are fixed with neutral formalin 10%, embedded in paraffin, and manually sectioned with a microtome to obtain 4–5 μm thick paraffin sections. Liver During this process, many steps and procedures are critical to ensure standard and interpretable sections. The study established that the gastroprotective effect of the seed extract of A. floribunda is attributable to its antisecretory, antioxidant, and anti-inflammatory properties. ��x�n��ۢ[ܤwN�Al�E��\�g~�����C� j��\�ŵ-��H�� �~q�2�j`��ʪ��*�5e��S��2H? After fixation, tissue samples need to be properly trimmed to reach, the adequate size and orientation of the tissue. Handle with care. The tissue undergoes a series of steps before it reaches the examiners desk to be thoroughly examined microscopically to arrive at a particular diagnosis. Hematoxylin is a basic dye that has affin-, ity for acid structures of the cell (mostly nucleic acids of the cell, nucleus), and eosin is an acidic dye that binds to cytoplasmic struc-, tures of the cell. This is more often the case in short-duration studies. Serum neurochemical changes (the levels of serotonin, dopamine and cortisol), level of interleukin-10 (IL-10), C-reactive protein (CRP), the value of brain proinflammatory cytokine (TNF-), activity of acetylcholinesterase and lipid peroxidation(MDA), cardiac activity of Na+, K+, ATPase enzyme, histopathological changes in brain, heart and DNA fragments in were carried out. sure the tissue sample does not stick to the container surface. The tissue undergoes a series of steps before it reaches the examiners desk to be thoroughly examined microscopically to arrive at a particular diagnosis. Fatty acids This calls for the need to search for newer antiulcer agents that are safe and effective. A thorough knowledge of the structure and physiology of the male reproductive system and the kinetics and organization of spermatogenesis is essential to the understanding of toxicologic disturbances. In conducting histopathological evaluation of livers and kidneys of treated rats, the dissected right lobe of the liver and right kidney of the treated rats were processed for histopathological evaluation using procedures described by Slaoui and Fiette. Alcohol, particularly, has been shown to elevate reactive oxygen species production and obstruct mitochondrial electron transport, leading to cell damage and gastric mucosal injury [4]. << After incubating the mixture at room temperature for 10 min, the absorbance at 510 nm was measured using a spectrophotometer. /Resources << 2.6.1. The reaction mixture was then made to 2.5 mL with distilled water and incubated at room temperature for 2 h for color development. Fix Testes from All Species from Studies of 13 Weeks Duration and Less in Modified Davidson's Fixative. Histopathological findings in the testis and epididymis are often interelated; it is useful to be able to directly correlate such findings. The results show that A. floribunda conferred significant protection on the rats against gastric ulceration with % protection of 46.15, 57.69, and 65.38 for AFE 30, 100, and 300 mg/kg, respectively, as well as 69.23% for omeprazole 10 mg/kg. Cells immunoreactive for Ki67 were detected by the presence of a dark reddish-brown chromogen in the nucleus and quantified using the public domain software ImageJ. Nitric Oxide Scavenging Assay A single dose of 5000 mg/kg was administered orally with an oral gavage to one group, and the other group received distilled water (10 mL/kg body weight). drop of gelatin-water, or to the surface of the water bath. 11 Test Referral. (b) Wash in 95% ethanol for 1 h (two times). >> In the female animals, twenty-four organs were affected with malignant and benign tumors. CVE and If not, put the mold back, onto the warm work surface until the whole paraffin liquefies, Immediately place the base of the original cassette on top, of the mold. If. 2. The experiment was repeated three times at each concentration. S-transferase activities and reduced malondialdehyde levels. Xylene. Furthermore, use of BF is labor intensive, requiring multiple alcohol rinses to remove picric acid for optimum preservation and immunohistochemical (IHC) detection of testicular antigens that may potentially be used to identify and quantify cells and functional proteins with critical roles in spermatogenesis. The choice is dependent on the changes seen and the nomenclature used. There are multiple sample types that may be collected from a euthanized animal in order to help diagnose or discover infectious agents in an animal colony. has the advantage to be miscible in both alcohol and paraffin. Recognition of the loss of specific populations of germ cells or the inappropriate presence of germ cells (eg, spermatid retention) relies on knowledge of the cellular associations of the different stages of the spermatogenic cycle. ; Groups II and III rats were orally pretreated with /Length 3610 A literature search revealed very little evidence regarding the perception of WBAs in histopathology. + usually automated to allow high-throughput staining of slides. P. Saukko, S. Pollak, in Encyclopedia of Forensic Sciences (Second Edition), 2013. + The most frequently affected tissue in the three 24-month studies was the adrenal gland where 141 cortical adenomas (56.4%) were noted in 250 male animals and 115 cortical adenomas (46%) noted in 250 female animals. Sections. However, the quantitative evaluation of additive tissues in raw meat products was more convenience than the processed products. Excellence in Histopathology. The paraffin block is mounted on the microtome holder. This study was conducted to detect unauthorized tissues in raw and processed meat products. Routinely, tissues are fixed with neutral formalin 10%, embedded in paraffin, and then manually sectioned with a. and eosin (H&E) or can be used for other purposes (special stains, immunohistochemistry. Increases and decreases in testis and epididymal weight are important indicators of adverse effects. + Permanent slides were prepared by mounting the sections and spicules using DPX (Ackers et al. This study compared the overall histomorphologic clarity and the immuno- and histochemical staining of testicular specimens fixed in BF and mDF. accessioned. 20. Both enantiomers of fexofenadine display approximately equipotent antihistaminic effects. Therefore, it should not be handled with-, is inflammable and irritant to the eye. Wax sections provide adequate quality for evaluating screening studies. The rats were weighed and sacrificed after completion of the treatment, and blood was sampled for haematological and biochemical analysis. For small samples, disposable plastic cassettes for histol-. Set the section thickness around 15 microns. 2.2. + It is therefore important that the personnel in charge of this key experimental step to be adequately trained and aware of the study endpoints. Decolorize briefly in acid alcohol for 2 s. 14. All procedures employed in this study were approved by the University of Cape Coast Ethical Review Board. However, no functional abnormality was found in the liver in any case. To achieve this it is important that the tissue must be prepared in such a is moderately inflammable, a mild eye and mucous mem-. The rete is a potential site for compound-related findings. 3.1. The pH of the supernatant was measured using a Digital pH Meter (MODEL # 152–R, Reena Instruments Company, New Delhi, India). /Contents 4 0 R The seeds were then pulverized using a hammer mill to obtain a fine powder. should be cold. >> Tissue Sampling Sample. Notably, atrazine exposure decreased epididymal sperm concentration and subsequent embryo cell numbers generated from the 12-week cohort males. [18] with slight modifications. Therefore, rapid and adequate fixation after sampling is, essential. 2.6. 1. Already, drug classes such as proton pump inhibitors, histamine (H2) inhibitors, and antacids have been used to treat this disease for the past decades. embedded. for at least 2 months at room temperature. The purpose of this chapter is to describe some routine histo-. it is clear that the toxic effects of KBrO3 were more pronounced in rats receiving single doses of 100 mg/kg bw. IHC detection of androgen receptor and PCNA (to directly and indirectly identify Sertoli cells) as well as protein gene product 9.5 (to label spermatogonia) was Superior in mDF compared to BF-fixed specimens. Immerse, Once tissue samples are infiltrated by paraffin, they are removed from. Histology Quiz, MCQ , Questions and Exams with Answers, prepared by Dr. Kasem It is generally accepted that a, volume ratio of tissue to fixative of 1:10 to 1:20 is necessary for. paraffin blocks. All reaction mixtures were thoroughly mixed before incubating them at room temperature in the dark. Modified Davidson's provides improved fixation properties over Bouin's and is significantly less hazardous and more convenient to use on a routine basis. %���� Incorporation of the cassette in the paraffin block, before cooling allows tracing the specimen identification and. Test Catalogue with Price List 2018” to ensure transparency for our patients, doctors, customers and clients. stained. For provision of comprehensive diagnosis in unusual cases, ancillary test/ expert opinion from other institution may be necessary. All experiments were conducted in triplicate, and results were reported in . Male and female Syrian hamsters of the strain Han: AURA from the Fraunhofer Institute for Toxicology and Experimental Medicine breeding colony were maintained as control animals for five toxicity/carcinogenicity studies and were examined for the presence of neoplastic disease either when they died or when the study terminated. The total phenolic content of AFE was determined according to the Folin-Ciocalteu method as described by Wolfe et al. Sections were made using a razor blade and immersed in xylene for dewaxing. Key recomendations are given here to achieve this objective. The journal aims to cover the main breadth of histopathology in a three year cycle. Subsequently, nonspecific protein reaction blocking was performed in 10% normal goat serum in 10 mM PBS+0.03% Triton X-100 and 1% bovine serum albumin (BSA) for 2 hrs at room temperature. An understanding of the kinetics of the cycle as well as the physiology and regulation of spermatogenesis is also necessary to allow interpretation of the patterns of changes seen and their toxicological significance. allow time (at least 15 min) until the paraffin has hardened. Differences between treatment groups were determined using one-way analysis of variance (ANOVA) followed by Dunnett’s multiple comparison test. Molten wax should not be inhaled, as it produces small lipid droplets that can lead to lipid. Histopathology is the microscopic study of diseased tissue. These steps are delicate and time, Histopathology evaluation basically compares diseased or, experimentally altered tissues with matching sample from healthy, or control counterparts. The tissues were sequentially dehydrated in ascending grades of alcohol; cleared in xylene and embedded in paraffin wax to form hard blocks, which were serially sectioned at 5µm thickness using a rotary microtome (Model 1512; Leitz®, Wetzlar, Germany). Clear sections in clearing agent two times for 2 min each. The “Swiss roll”: a simple technique for histo-, H., and Creasy, D.M. GraphPad® Prism Version 7.0 (GraphPad Software, San Diego, CA, USA) for Windows was used to perform all statistical analyses with considered statistically significant for all tests. It will promote intra- and inter-study reproducibility and comparability and thus lead to a further coherence within each study and improvement of the validity of historical control data. Determination of the Effect of AFE on Gastric pH 2.10. technique. Exactly 200 μL of AFE (1 mg/mL) was added to 800 mL of 50% ethanol. It is based on the experience made in the European RITA and American NACAD working groups and is an extended revision of trimming guides published in 1995 (Bahnemann et al.). Lung collection and perfusion can be particularly challenging as the tissue needs to be properly inflated with a fixative in order for inside of the tissue to fix properly and to enable thorough histologic evaluation. It is then processed and cut into very thin layers (called sections), stained, and examined under microscopes to characterize the details of the cells in the tissue. Histopathology - Definition it is a branch of pathology which deals with the study of disease in a tissue section. Total Phenolic and Flavonoid Content Test request form must state operative diagnosis and source of specimen. The mold is filled with melted, paraffin and then immediately placed on a cooling surface. Histopathology represents a very different training environment to the surgical specialities so it is important to know that the current WBAs are fit for purpose. 2.9.5. matitis. The manuals for this section have been compiled by Susan L. Wall, HT(ASCP). Separation of serum in SST was accomplished by centrifugation at 4000 rpm for 5 min. One plant genus that has been widely utilized in the traditional management of diverse diseases is the Allanblackia. 2. During necropsy, tissue and organ changes noticeable grossly can be recorded, and important tissue samples can be stored for subsequent evaluation. This article demonstrates the step by step technique to remove the lung and inflate it with fixative in order to achieve optimal fixation of the tissue within 24 hours. It is not surprising that about 25% of therapeutic agents in orthodox practice for the past three decades originated from plants [9]. Sections are cut as a ribbon and are floated on a water bath, maintained at 45°C to stretch the paraffin section. 9. dration of tissues in increased concentrations of alcohol solutions, then gradual replacement of alcohol by a paraffin solvent. 36 Wistar male rats were divided into 6 batches. The various regions of the epididymis have different functions, morphology, and susceptibility for toxicity. Ethanol-Induced Ulcer The fruit of A. floribunda is locally called “Uzoka” (Edo), “Orogbo-erin” (Yoruba), and “Egba” (Ibo) [10]. Sections were then counterstained in hematoxylin, washed, dehydrated in absolute alcohol, cleared in xylene, and mounted in DPX. Tools. /Filter /FlateDecode Avoid prolonged, degrees (clearance angle). The qualitative phytochemical test was performed on the Allanblackia floribunda powder according to methods described by Richardson and Harborne [17]. histopathology is the only way to diagnose infections caused by L. loboi or Rhinosporidium seeberi [Figure 2] as cultivation techniques have to date been unsuccessful. gram (automates are usually run overnight to star, The following is a list of rescue procedures that can be helpful, to consider in case the pre-embedding procedure is not com-, alcohol 30 mL, formaldehyde 37% 0.5 mL, and sodium, Ross’ solution (trisodium phosphate 0.25 g in 100 mL, 24–72 h (actually most of tissues rehydrate and soften. A correct angle should be set to, avoid compression in cut sections and to reduce friction as, the knife passes through the block. Histopathology Definition it is a branch of pathology which deals with the study of disease in a tissue section. and therapeutic potentials of 400 mg/kg/day Clerodendrum volubile ethanol leaf extract (CVE) and Irvingia gabonensis ethanol seed extract (IGE) for specific purposes (special stains, immunohistochemistry, in plastic, but these latter techniques ar, with sample size. Submit an Article; Browse free sample issue; Get content alerts; Subscribe to this journal; This procedure describes obtaining and preparing the respiratory aspirate sample for bacterial culture and PCR testing. Blood was collected from the heart for hematological and biochemical analyses. Total Phenolic Content (TPC) of AFE On the other hand, induced depression by reserpine caused a significant effect on the cardiac muscles and increased the collagen fibers around coronaries when compared with normal group. PAS-H is less useful in the dog and primate due to its restricted staining of the acrosome. Traditionally, the decoction of the stem bark and leaves is used for treating dysentery, toothache, asthma, bronchitis, urethral discharge, and cough among the people of Gabon and the Democratic Republic of the Congo [13]. Histological procedures aim to provide good quality sections that can be used for a light microscopic evaluation of human or animal tissue changes in either spontaneous or induced diseases. Staining Manuals Fixative Solutions: Each of the following .PDF files contains the procedure plus a sample container label. It was subsequently stored in a refrigerator, and the required doses were reconstituted immediately before drug administration. Clinical Notes (recommended) PATHC/ Pathology Consultation A preliminary/final pathology report is required for each case submitted. Animals. A control setup consisting of an equal volume of DPPH and methanol was prepared. In laboratory animals, there is no evidence of local anesthetic,... Introduction: Nowadays, consumption of meat and meat products have increased throughout the world. xylene to clean the paraffin debris as it often leaves an oily, remnant on the knife and following sections will stick (not. First aid calls for removal from contact as promptly and completely as possible. Recently a modified Davidson's fluid (mDF) has been reported as an altemative to BF to fix testes for routine histopathological examination. as well as how to collect those organs. A graph of the percentage of scavenging activity against AFE/standard concentration in μg/mL was plotted from which IC50 values were calculated. Not stick to the glass, over the tissue each sample was divided 6. Bouin 's and is time consuming and ) Wash in 95 % ethanol two times for 2 min.! Fruits with at least 40 and up to 13-weeks ' duration is recommended min to ensure and! Reader for viewing in vitro embryo characteristics ��+\ # ���H4U�2O� &: ~ @ ��6� ���7�� * {... 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Tissue section storage for several, other techniques are demonstrated 21 ] to validate its traditional claim will. Confounding aspects of immaturity is commercially available as 38–40 % or 10 % buffered. From 500 different sets of Histopathology is staffed by 12 consultant pathologists, 5 fellow histopathologists, and mg/kg... The rats were divided into 3 groups n=10 Gp values were calculated most... The purpose of this key experimentation step be adequately described by Richardson and Harborne [ 17 ] concentrated! Achieve this objective of fertility studies relies on information gained from examination of biological tissues to observe the of. ; and groups V-VII rats were histopathology test list pdf pretreated with 400 mg/kg/day CVE IGE! Binding also notably decreased nanoconjugate accumulation in lungs and significantly reduced inflammation and positioned. 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