Routine Staining Research Articles

Oncogenic Alterations in Histologically Negative Lymph Nodes Are Associated with Prognosis of Patients with Stage I Lung Adenocarcinoma.

Simple SummaryLymph nodes (LNs) metastasis is one of the most important factors affecting the outcome of non-small cell lung. The aim of this study is to explore whether presence of oncogenic alterations in histologically-negative lymph nodes (LNs) can be of prognostic significance in stage I lung adenocarcinoma (LUAD). We confirmed that presence of oncogenic alterations in regional LN may be associated with higher risks of postsurgical recurrence of Stage I LUAD, particularly for certain molecular subgroups. These results warranted future studies on larger cohort of NSCLC patients using more comprehensive cancer gene panels to establish the clinical impact of molecular LN occult metastasis for localized NSCLC and identify Stage I patients at high risks for recurrence for appropriate adjuvant therapy.Background: Survival of patients with stage I non-small cell lung cancer (NSCLC) varies greatly. We sought to explore whether presence of oncogenic alterations in histologically-negative lymph nodes (LNs) can be of prognostic significance in stage I lung adenocarcinoma (LUAD). Methods: Genomic analysis of oncogenic alterations was applied to 123 stage I LUAD tumors. The same genomic variants identified in primary tumors were examined in corresponding histologically-negative LNs. Results: A total of 102 (82.9%) patients had at least one canonical oncogenic alteration detected in primary tumors, and 57 LNs from 12 patients (11.8%) were found to carry the identical oncogenic alterations detected in the corresponding primary tumor tissues, including EGFR mutations (six cases), KRAS mutations (three cases), ALK fusion (one case), BRAF mutation (one case) and HER2 & NRAS co-mutations (one case). None of these LNs was found to have occult tumor cells by routine pathological assessment or immunohistochemistry staining using antibodies against pan-cytokeratins (AE1/AE3) and the epithelial marker Ber-EP4. The detection rate of oncogenenic alterations in LN was significantly higher in RAS-mutant tumors than EGFR mutant tumors (36.36% verse 7.41%, p = 0.017). Patients with oncogenic alterations in LN showed inferior disease-free survival (DFS, p = 0.025) and overall survival (OS, p = 0.027). Furthermore, patients with RAS-mutations detected in LN had the worst DFS and OS (p = 0.001). Among the 11 patients with RAS mutation in primary tumors, DFS and OS in the four patients with mutations detected in LN were significantly shorter than the remaining seven patients without mutations LN (DFS, p = 0.001, OS, p = 0.002). Conclusions: Genomic analysis has the potential to detect oncogenic alterations in regional LNs for localized LUAD and presence of oncogenic alterations in regional LN may be associated with inferior clinical outcome of stage I LUAD, particularly for certain molecular subgroups. ClinicalTrials.gov ID NCT04266691

Unveiling idiopathic guttate hypomelanosis: pathology, immunohistochemistry, and ultrastructural study.

AbstractBackgroundIdiopathic guttate hypomelanosis (IGH) is a pigment disorder of unknown etiology. Despite its high prevalence and the unaesthetic appearance of the lesions, there are relatively few histological studies on this disorder. This is an important gap to understanding its pathogenesis.ObjectivesTo assess the microscopic structure of IGH lesions compared to normal adjacent skin areas and the possible interaction between melanocytes and the subjacent dermis.MethodsIn this cross‐sectional study, we took biopsy specimens of hypochromic lesions and adjacent normal skin from 20 patients with IGH. We analyzed the fragments using routine stains, immunohistochemistry, and electron microscopy.ResultsWe found superficial dermal fibrosis in 90% (18/20) of our IGH cases and unreported keratinocyte cytoplasmic changes on electron microscopy.ConclusionOur results suggest an interaction between melanocytes and the subjacent dermis in IGH. These findings can help to understand melanocyte biology and the pathogenesis of other achromic lesions.

MRNA expression of the severe acute respiratory syndrome-coronavirus 2 angiotensin-converting enzyme 2 receptor in the lung tissue of Wistar rats according to age.

Background and Aim:Angiotensin-converting enzyme 2 (ACE2) is expressed and plays functional and physiological roles in different tissues of the body. This study aimed to distinguish the levels of expression of ACE2 in the lung tissue at different ages of rats.Materials and Methods:In this study, 18 male rats were used and divided into three groups according to age. Real-time quantitative polymerase chain reaction (RT-qPCR) was conducted to determine the levels of the quantification of eosinophil cationic protein mRNA transcript. In addition, tissue specimens of the lung were stained with routine hematoxylin and eosin stains.Results:This study confirmed that RT-qPCR amplification plots of ACE2 gene exhibited clearly expression of the lung tissue of rats in the different groups and there are strong different threshold cycles numbers according to the age at 2 weeks, 2 months, and 6-8 months. Consequently, the expression of ACE2 was completely different between groups depending on the age of the rats. The RT-qPCR results showed that the older animal group (age of 6-8 months) had a significantly higher expression of ACE2 than the other animal groups (ages of 2 weeks and 2 months). In the same way, the second group (age of 2 months) had a significantly higher expression of ACE2 than the first group (age of 2 weeks). This study confirmed that the ACE2 expression is influenced by the age of rats.Conclusion:This study concluded that the expression of the ACE2 receptor of coronavirus disease 2019 would be different according to the age of rats, and this result suggested that expression of ACE2 in lung tissue could determine infection and pathogenesis of COVID-19 during different ages of rats or some individual differences.

Beetroot and Turmeric as Alternative Dyes for Haematoxylin and Eosin in Histological Staining

Dyes are referred to as stains which are used to add color to tissues and microbes to make them optical distinct. The dyes are both synthetic and natural, the synthetic ones may be harmful, hazardous to health and are costly to obtain. Natural dyes are eco-friendly, not costly and not hazardous to health as compared to synthetic dyes. In this study, various procedures such as H and E which serves as control as normal routine staining in histopathology laboratory was used to compare Beetroot and Eosin (B and E), Haematoxylin and Turmeric (H and T). Beetroot and Turmeric (B T) Beetroot and Eosin stains collagen fibre almost similar to H and E, that shows beetroot can be used as alternative for Haematoxylin in H and E staining technique. Haematoxylin and Turmeric stained the tissue almost similar to H and E which shows that Turmeric can replace Eosin. However, B and T poorly stained collagen fibre when compared with H and E. The conclusion came out that, Beetroot containing Betanin which gives it a red color has the same ability of staining Nuclei Bluish purple because of its basic content and may require a mordant (Vineger) as attain enhancer for most of the tissues. Turmeric exhibits the basophilic-like effect on collage fibre, it stains the cytoplasm fairly yellowish, because of its curcumin that makes it yellow. Beetroot and Turmeric as a direct replacement of H and E staining technique in Histopathology laboratory should further be studied with adequate concentration and mordant as the features is suggestive that B and T can replace H and E.

CmyoSize: An ImageJ macro for automated analysis of cardiomyocyte size in images of routine histology staining

BackgroundCardiomyocyte size histomorphometry is widely required in myocardial remodeling research, but it relies on subjective, time-consuming manual tracing. We aimed to present the CmyoSize, a Fiji/ImageJ macro that reproducibly measures the transnuclear cross-sectional size of cardiomyocytes in multiple H&E images. MethodsCmyoSize output per image are: mean cross-sectional area (CSA), CSA’s local standard deviation (LSD), mean width, and width’s LSD. CmyoSize was validated against measurements of an expert examiner, using myocardium micrographs (400-fold magnification) from rats treated with hyperbaric oxygen (HBO), streptozotocin (STZ), as well as control rats (five images/heart, five rats/group). ResultsCmyoSize achieved high cardiomyocyte detection precision (95.17 ± 1.11%) and it analyzed each image at least 30-times faster than expert examiner (8.44 ± 0.17 s vs. 266.70 ± 25.15 s). Bland-Altman analyses showed no meaningful bias ( ± 95% confidence interval) between both methods: mean CSA = 1.88 ± 6.83 µm2, CSA’s LSD = −4.55 ± 5.07 µm2; mean width = −0.02 ± 0.22 µm; width’s LSD = −0.16 ± 0.17 µm. Pearson’s r ranged from 0.85 to 0.94 (P < 0.0001). CmyoSize revealed homogenous cardiomyocyte hypertrophy in HBO-treated animals and STZ-induced heterogeneous hypertrophy. ConclusionFully automated, standardized cardiomyocyte size quantification in H&E images using CmyoSize is feasible, accurate, and time-efficient.

Growth factors and chromogranin A in intra-abdominal adhesions of children

Pathogenesis of adhesions is still unclear and does not give a clear response whether the morphopathogenetic mechanisms of congenital and acquired adhesions in children are different or not. Thus, we searched for the appearance and distribution of growth factors, their receptors and chromogranins in adhesions of children in the ontogenetic aspect. Tissues were obtained from 17 children of group 1 (1 day to 3 years old) and 10 children of group 2 (10 to 16 years old). Visibly intact peritoneum was obtained from teenagers of group 2 and used as control tissue. Routine staining and immunohistochemistry for insulin-like growth factor 1 (IGF1), insulin-like growth factor 1 receptor (IGF IR), transforming growth factor beta (TGF-β), hepatocyte growth factor (HGF), chromogranin A (CgA) was applied. Statistical analysis included semi-quantitative evaluation of immunopositive structures, Wilcoxon signed-rank test and Wilcoxon rank sum test for comparison of data. The results showed patchy fibrosis with round-shaped fibroblasts, neoangiogenesis and inflammation. Adhesions in group 1 demonstrated various appearance of IGF, IGF IR, few to moderate number of TGF-β-, HGF- and moderate number of CgA-containing cells. In both groups, fibroblasts predominated among actively expressing cells. Group 2 showed moderate to numerous IGFI and HGF positive cells, while IGF IR- and TGF-β-containing cells were numerous. Intact peritoneum showed variable TGF-β, CgA, and few to moderate HGF, IGFI, IGF IR cells. In conclusion, an increase in IGFI, IGF IR, TGF-β and HGF characterises the acquired (“aged”) adhesions. Fibroblasts are most active in adhesion formation with the increase of cellular activity during the aging of children and adhesions. Fibroblasts with changes in shape stimulate further development of disease via intensive CgA expression.

Histopathological Study of Experimentally Crushed Skeletal Muscle’s Regeneration in Adult Albino Rats

Aim: This study aims to explain all the events of skeletal muscle repair and regeneration with the help of suitable histophathlogical photomicrographs taken from crush-injured adult albino rat’s gluteus maximus muscle. Materials and Methods: The present study is part of our previous research study related to skeletal muscle repair and regeneration in crush injured gluteus maximus muscle of adult albino rats. The samples were processed for histopathological examination using routine and special histological staining procedures. The tissue samples were examined under trinocular microscope, and the fields showing interesting findings were recorded under different magnification. Results: In this study we observed all regenerative changes in myofibers and related structures after crushed injury. Conclusion: Histopathological studies with good stainings are helpful for the easy identification of minute changes that occurs in each stages of skeletal muscle regeneration.

Evaluation of Multiplex Real-time PCR and WHO Criteria for Diagnosing Childhood Bacterial Meningitis in a Tertiary Referral Hospital in Iran

Background: Childhood bacterial meningitis (BM) requires prompt and precise diagnosis to provide proper treatment and decline mortality and morbidity. Objectives: We aimed to evaluate the World Health Organization (WHO) criteria and polymerase chain reaction (PCR) for diagnosing BM in children admitted to a tertiary referral hospital in Shiraz, southern Iran. Materials: We included all 492 children aged one month to 17 years suspected of meningitis who had cerebrospinal fluid (CSF) leukocytosis admitted to Nemazi Hospital from August 2016 to September 2017. The CSF specimens were examined for routine analysis, Gram staining, and culture. A multiplex real-time PCR was used to identify Streptococcus pneumoniae, Haemophilus influenzae type b (Hib), and Neisseria meningitidis in the CSF samples. Seven viruses were also investigated using real-time PCR. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were calculated using the WHO criteria and the multiplex real-time PCR results. Results: Seventy-four CSF samples had leukocytosis. Nineteen (22.9%) patients had BM caused by S. pneumoniae (n = 14), Hib (n = 2), Salmonella enterica (n = 2), and N. meningitidis (n = 1). The PCR test detected all cases, except for two with Salmonella meningitis (sensitivity 89.4%, specificity 100%, PPV 100%, and NPV 96%). The WHO criteria detected all cases, except three who received antibiotics at least four days before performing lumbar puncture (sensitivity 84.2%, specificity 98.2%, PPV 94.1%, and NPV 94.7%). Enterovirus was the most common viral etiology (6.75%). Conclusions: The WHO criteria and the multiplex real-time PCR had high accuracy in our setting, and their use could decrease the antibiotic over-prescription in febrile children suspected of meningitis.

Morphofunctional characteristics of the heart in sudden death of children under 1 year of age in the forensic medical aspect

The aim of this study is to describe the morphological and functional characteristics of the cardiac in sudden infants death syndrome (SIDS) in children under one year of age. Twenty eight cases of SIDS were studied histologically and immunohistochemically. Histological examination of the sectional material was carried out using standard and additional stains. The study of cardiac muscle tissue with routine staining with haematoxylin-eosin revealed interstitial oedema, uneven plethora of blood vessels: dystonia and weak plethora of part of the coronary arteries, excess of red blood cells in the veins as well as microcirculatory plethora with stasis of the erythrocytes. An immunohistochemical study revealed a mild over expression of p53 in cellular elements, small-focal expression of CD68 in cardiomyocytes apoptosis intramural areas, activation of mast cells (CD117), expression of ki-67 in macrophages, proliferation of fibroblasts. Additional forensic criteria for the diagnosis of SIDS were determined in the form of atrial endocardial fibrosis and interventricular septum; the expression of CD68, CD117 in fibroblasts, mast cells and lymphocytes; apoptosis of cardiomyocytes (expression of p53), proliferation of fibroblasts and remodeling of the heart (expression of ki-67).

The clinical, pathological, and genetic characteristics of lipid storage myopathy in northern China.

The lipid storage myopathy (LSM) diagnosis is based on the patient's clinical manifestations and muscle pathology. However, when genetic testing is lacking, there is a high rate of misdiagnosis of the disease. This study aimed to investigate the clinical and pathological features of genetically diagnosed LSM in northern China, analyze genetic mutations' characteristics, and improve the LSM diagnostic rate. Twenty patients with LSM diagnosed were collected; meanwhile, the clinical data, muscle samples, and routine pathological staining of muscle specimens were collected. The morphological changes of muscle fibers were observed under an optical microscope. Among the included patients, 18 cases had ETFDH (HGNC ID: 3483) mutations, and two had PNPLA2 mutations. Family pedigree verification was performed on three patients with heterozygous mutations in the ETFDH gene complex. Histopathological staining showed that all patients had fine vacuoles in the muscle fibers, and some of them merged to form fissures, and the lipid droplets increased in cells. After therapy, 18 patients were associated with a favorable prognosis, and two patients were ineffective with the treatment of neutral lipid storage myopathy (NLSDM) caused by PNPLA2 mutation. The clinical manifestations of LSM are complex and diverse, mainly manifested by proximal muscle weakness and exercise intolerance in the extremities. The pathological images of LSM muscles are abnormal storage of lipid droplets in muscle fibers, primarily involving type I fibers. The LSM patients were mainly multiple acyl-CoA dehydrogenase deficiency (MADD) caused by the ETFDH gene mutation. It is necessary to perform an accurate typing diagnosis of LSM.

Upregulation of C/EBP Homologous Protein induced by ER Stress Mediates Epithelial to Myofibroblast Transformation in ADTKD-UMOD.

Autosomal dominant tubulointerstitial kidney disease due to UMOD mutations (ADTKD-UMOD) results in chronic interstitial nephritis, which gradually develops into end-stage renal disease. It is believed that the accumulation of mutant uromodulin causes the endoplasmic reticulum (ER) stress, then leads to the kidney damage. But the underlying mechanism remains unclear. To find the ADTKD-UMOD patients, UMOD gene screening was performed in 26 patients with unexplained chronic interstitial nephritis, during the past 10 years in our department, and among them three ADTKD-UMOD cases were discovered. Routine pathological staining and electron microscopy sections were reviewed again to confirm their kidney lesions. Immunostaining of UMOD and ER stress marker GRP78, as well as CHOP have all been done. The strong colocalization of UMOD with GRP78 and CHOP in ADTKD-UMOD patients but not in other chronic interstitial nephritis patients had been found. Moreover in vitro experiments, ER stress induced by tunicamycin (TM) not only significantly increased the expression of GRP78 and CHOP, but also caused the epithelial to myofibroblast transformation (EMT) of renal tubular epithelial cells, evidenced by decreased expression of E-cadherin and increased expression of vimentin, and extracellular matrix (ECM) deposition, evidenced by increased expression of fibronectin (FN). CHOP knockdown could restore the upregulation of vimentin and FN induced by TM. Thus, specific activation of CHOP in renal tubular epithelial cells induced by UMOD protein might be the key reason of renal interstitial fibrosis in ADTKD-UMOD patients.

Findings of limb-girdle muscular dystrophy R7 telethonin-related patients from a Chinese neuromuscular center.

Limb-girdle muscular dystrophy (LGMD) is a group of clinically and genetically heterogeneous neuromuscular disorders. LGMD-R7, which is caused by telethonin gene (TCAP) mutations, is one of the rarest forms of LGMD, and only a small number of LGMD-R7 cases have been described and mostly include patients from Brazil. A total of two LGMD-R7 patients were enrolled at a Chinese neuromuscular center. Demographic and clinical data were collected. Laboratory investigations and electromyography were performed. Routine and immunohistochemistry staining of muscle specimens was performed, and a next-generation sequencing panel array for genes associated with hereditary neuromuscular disorders was used for analysis. The patients exhibited predominant muscle weakness. Electromyography revealed myopathic changes. The muscle biopsy showed myopathic features, such as increased fiber size variation, muscle fiber atrophy and regeneration, slight hyperplasia of the connective tissue, and disarray of the myofibrillar network. Two patients were confirmed to have mutations in the open reading frame of TCAP by next-generation sequencing. One patient had compound heterozygous mutations, and the other patient harbored a novel homozygous mutation. Western blotting analysis of the skeletal muscle lysate confirmed the absence of telethonin in the patients. We described two LGMD-R7 patients presenting a classical LGMD phenotype and a novel homozygous TCAP mutation. Our research expands the spectrum of LGMD-R7 due to TCAP mutations based on patients from a Chinese neuromuscular center.

Technical Note: Measuring the thickness of histological sections by detecting fluorescence intensity of embedding foam

Fluorescence intensity of embedding foam in paraffin blocks can be used to measure the thickness of histological microsections. We embedded samples of embedding foam and produced several microsections of varying thicknesses using routine processing and staining. Fluorescence intensity in the blue area of the embedding foam detected with a slide scanner was compared to absolute thickness as measured using confocal microscopy. Correlation analysis displayed a clear linear correlation with convincingly low prediction interval. The concept of measuring thickness of histological microsections by detecting fluorescence intensity of embedding foam is suggested as an approach to high-throughput measuring of histological sections applicable for a fully digitized pathology department. No acquisition of dedicated equipment is required and the method can be applied as a fully automated technique requiring no time consumption.

Surge and Scare of Mucormycosis in the Backdrop of COVID-19 Pandemic

Introduction: Mucormycosis is a rare opportunistic fungal infection. During the second wave of Coronavirus Disease 2019 (COVID-19), a remarkable increase in mucormycosis cases was observed. Diabetes mellitus is an independent risk factor for both mucormycosis and COVID-19. Aim: To assess the incidence of mucormycosis cases in COVID19 infected and non COVID-19 infected patients and also to assess the related co-morbid conditions during the second wave of COVID-19 pandemic. Materials and Methods: This cross-sectional study was conducted from 15th May 2021 to 15th July 2021 in the Department of Pathology, GGH/Government Medical College, Srikakulam, Andhra Pradesh, India. The study included 63 cases of mucor suspects. The parameters analysed in this study were age, sex, COVID-19 status, co-morbidities, steroid status and radiological findings. A descriptive analysis was made from the data obtained. The information was analysed in data spreadsheets using Microsoft office excel 2010. Results: Out of total 63 mucor suspects, 60 (95%) cases tested positive for COVID-19 and 44 (73.3%) cases of them received corticosteroid treatment. Mean age group of 41-50 years with 27 (42.8%) cases with a male preponderance of 49 (77.7%) cases was noted. The most common identified risk factor was diabetes mellitus seen in 22 (34.9%) cases and diabetes associated with hypertension in 14 (22.2%) of the cases. Mucormycosis fungal hyphae were noted in 77.7% (49) of cases on routine histopathological examination stain and 74.6% (47) of the cases by special stains for confirmation. Conclusion: In the present study, it was observed that mucormycosis infection was most commonly seen in COVID19 infected patients in post-COVID-19 phase and also in minority of non COVID-19 infected cases with prior debilitating conditions. Therefore, it was concluded that, mucormycosis was positively associated with COVID-19 infected patients who have co-morbidities like diabetes mellitus and history of steroid usage.

Ex situ perfusion fixation for brain banking: a technical report.

Perfusion fixation is a well-established technique in animal research to improve preservation quality in the study of many tissues, including the brain. There is a growing interest in using perfusion to fix postmortem human brain tissue to achieve the highest fidelity preservation for downstream high-resolution morphomolecular brain mapping studies. Numerous practical barriers arise when applying perfusion fixation in brain banking settings, including the large mass of the organ, degradation of vascular integrity and patency prior to the start of the procedure, and differing investigator goals sometimes necessitating part of the brain to be frozen. As a result, there is a critical need to establish a perfusion fixation procedure in brain banking that is flexible and scalable. This technical report describes our approach to developing an ex situ perfusion fixation protocol. We discuss the challenges encountered and lessons learned while implementing this procedure. Routine morphological staining and RNA in situ hybridization data show that the perfused brains have well-preserved tissue cytoarchitecture and intact biomolecular signal. However, it remains uncertain whether this procedure leads to improved histology quality compared to immersion fixation. Additionally, ex vivo magnetic resonance imaging (MRI) data suggest that the perfusion fixation protocol may introduce imaging artifacts in the form of air bubbles in the vasculature. We conclude with further research directions to investigate the use of perfusion fixation as a rigorous and reproducible alternative to immersion fixation for the preparation of postmortem human brains.

Evaluation of the efficacy of hematoxylin and eosin stain when xylene is completely replaced by turpentine or kerosene oil - A comparative study for oral tissues.

Microscopic examination of cells and tissues requires the preparation of very thin and good-quality sections mounted on glass slides and appropriately stained to demonstrate normal and abnormal structures. Before this step, the tissue must undergo preparatory treatment known as tissue processing. The various stages of tissue processing are dehydration, clearing, impregnation, and embedding, each with a particular duration for proper completion of the process. Xylene is the most frequently used clearing agent whose carcinogenic potential is well documented. Hence, attempts were made to substitute xylene with a biosafe clearing agent. The present study aimed to evaluate and compare the efficacy of hematoxylin and eosin stain (H and E stain) when xylene is completely replaced by turpentine or kerosene oil. A total number of 50 tissue samples were taken in the study, which included 40 study samples and 10 controls. All the samples were randomly separated into three groups and routine tissue processing and H and E staining were performed. The result was further subjected to statistical analysis by using Fisher's exact test. Group-1: Ten tissue samples were processed and H and E staining was done in xylene. Group-2: Twenty tissue samples were processed and H and E staining was done in turpentine oil. Group-3: Twenty tissue samples were processed and H and E staining was done in kerosene oil. Nuclear staining, cell morphology, and uniformity of staining were better in kerosene sections, while cytoplasmic and clarity of staining of turpentine sections were comparable with xylene sections. Turpentine and kerosene as clearing agents can be used in the future with certain modifications in their concentration and routine staining protocol.

Pleural pathology in HIV-infected patients: features of morphological diagnostics

Background. The progression of HIV infection is accompanied by the development of opportunistic diseases, including pleural effusions of various origins. Morphological examination of pleura tissue in cases of pleural effusion serves as the basis for establishing the etiology of the pathological process and, therefore, the final clinical diagnosis. Objective – analysis of results of morphological diagnostics of pleura lesions in HIV-infected patients in comparison with other laboratory tests and clinical diagnosis. Methods. 103 cases of pleurisy of various origins were studied. Pleural biopsies were obtained by various types of minimally invasive diagnostic interventions with subsequent morphological examination. Results were compared with the available data of microbiological and molecular genetic studies of pleural biopsies. Results. At the time of hospitalization the preliminary clinical diagnosis was pleurisy of unknown etiology in 96,1 % of cases. A combined disease was diagnosed – hepatitis C in a third of all observations according to the results of laboratory tests. Due to routine staining with hematoxylin and eosin, tuberculosis lesions of the pleura were diagnosed in 59,2 %, the second most frequent was the diagnosis of nonspecific pleurisy, 20,4 %. According to the duration of the process, acute pleural tuberculosis was established in 19,7 % of cases, the subacute form of tuberculosis pleurisy – in 54,1 % of cases, and chronic pleural tuberculosis was established in 22,9 % of cases. In 17,5 % of cases, in order to clarify the etiology of pleurisy, additional histochemical staining for infectious agents was performed. The results of microbiological and molecular genetic studies were established in 76,7 % of cases. The greatest number of M. tuberculosis detection was obtained during the culture study of the biopsy material and exudates. When comparing the final clinical diagnosis and the level of CD4 cells in peripheral blood, it was found that in most cases (74,5 %) pleural effusions developed at low counts of CD4 cells, less than 350/l. Conclusion. Tuberculosis predominates in the etiological structure of pleural effusions in patients with HIV infection. Pleural tuberculosis can be the main secondary disease or be combined with pulmonary tuberculosis. In second place in terms of frequency of occurrence, nonspecific pleurisy was diagnosed as a complication of the main secondary disease. Pleural effusions develop when CD4 cell counts are low. Morphological diagnostics of pleural lesions is the main research method in the diagnostic algorithm of cases of pleural effusions of unknown etiology against the background of HIV infection.

Utility of nail enamel for inking of surgical margins in oral biopsy: a comparative study

Background: India Ink is the most commonly used ink in surgical pathology. The main disadvantages of India Ink are longer drying time, monochrome, toxicity and cost. Because of these disadvantages, alternative materials have been suggested to replace India ink. The aim of this study is to evaluate the effectiveness of nail enamel for inking of surgical margin and to compare it with India ink. Materials and methods: N = 20, which included 10 mucosal and 10 skin samples. Each selected margin is divided into 2 equal halves and one is inked with India ink and the other with nail enamel (Vernis A Ongles: Dark green). After routine processing and staining, the effectiveness of nail enamel and India ink were compared based on macroscopic and microscopic parameters. Results: Less drying time and visibility on paraffin wax block were excellent for nail enamel. Microscopic visibility of nail enamel was comparable with that of India ink. However, processing fluids contamination is the main drawback of nail enamel. Conclusion: Nail enamel can be used as an alternative to India ink because of its less drying time, ease of application, good visibility on wax blocks and microscopically, availability in multiple colours, cost effectiveness and non-toxicity.

2021 Canadian Surgery Forum: Virtual, online Sept. 21-24, 2021.

# 01. Design and validation of a unique endoscopy simulator using a commercial video game {#article-title-2} Procedural simulation has been shown to enhance early endoscopy training. In this proof of concept study, we aimed to show that a first-person shooter (FPS) video game with a novel in-house

Standard techniques for formalin-fixed paraffin-embedded tissue: A Pathologist’s perspective

Histopathology considered as an essential keystone to understanding diseases on a cellular level, without examining affected tissues and cells; will lack the accurate diagnosis. The most common protocol that used worldwide for histopathology is termed as formalin-fixed paraffin-embedded techniques; this protocol is the oldest protocol and still used, this technique not invented once, but it is started with a simple process of tissue fixation then undergo several successful steps as it's known today. This protocol depends on formalin to induce tissue fixation and preservation simultaneously, followed by a series of steps to facilitate infiltration of hot paraffin wax in the cells to adjust the density of tissue similar to that of paraffin wax to support tissue during sectioning process at few micrometers of thickness. Many protocols have been invented and modified subsequently to increase the accuracy of tissue processing and facilitate their study under light microscopy. Additionally, the formalin-fixed and paraffin-embedded samples can be considered the most durable process till now. In this review, we are trying to view all fundamental aspects of classical tissue processing that was depended since 1850s, starting from tissue examination, sampling, accessory, labelling, fixation, dehydration, clearance, infiltration, and embedding with hot paraffin wax. Additionally, explain the routine sectioning and staining with classical Hari’s hematoxylin and alcoholic eosin staining protocol.