Nuclear Details Research Articles

Evaluation of efficacy of commercially available decalcifying agents

Aims and Objective: Decalcification of hard tissues specimens for the purpose of histopathological examination is often a time-consuming process and lead to undue delay in diagnosis. As a part of search for an ideal decalcifying agent, we designed the present study to compare two commercial decalcifying agents with 5% nitric acid Materials and Methods: Size matched bone specimens from human mandible were decalcified, using 5% Nitric acid, commercial decalcifying agents: Product A and Product B (n=22 each). Decalcified specimens were processed, sectioned, stained with H&E stain and evaluated for different parameters such as microtomy, staining quality, structural integrity, preservation of cellularity and cellular and nuclear details. The results were compared between the groups using Krunskal Wallis test and Mann-Whitney U test. Results: 5% Nitric acid is found to be significantly better in terms of microtomy, staining characteristics and structural integrity with p-value being 0.002, 0.005 and 0.004 respectively when compared to Product A. Similarly microtomy, staining characteristics and preservation of cellularity were noted to be superior with 5% Nitric acid in comparison with Product B, with p-value of 0.004, 0.001 and 0.019 respectively. None of the assessed parameters showed significant difference between Product A and Product B groups. Conclusion: We found 5% nitric acid to be a better decalcifying agent compared to the commercial decalcifying agents used in this study. When time is a constrain, Product A i.e Osteomoll is a good choice, as the process of decalcification is faster without compromising the quality to a great extent. Keywords: Decalcification, 5% Nitric acid, Commercial decalcifying agents.

Neurosurgical Flexible Probe Microscopy with Enhanced Architectural and Cytological Detail

Microscopic delineation and clearance of tumor cells at neurosurgical excision margins potentially reduce tumor recurrence and increase patient survival. Probe-based invivo fluorescence microscopy technologies are promising for neurosurgical invivo microscopy. We sought to demonstrate a flexible fiberoptic epifluorescence microscope capable of enhanced architectural and cytological imaging for invivo microscopy during neurosurgical procedures. Eighteen specimens were procured from neurosurgical procedures. These specimens were stained with acridine orange and imaged with a 3-dimensional (3D)-printed epifluorescent microscope that incorporates a flexible fiberoptic probe. Still images and video sequence frames were processed using frame alignment, signal projection, and pseudo-coloring, resulting in resolution enhancement and an increased field of view. Images produced displayed good nuclear contrast and architectural detail. Grade 1 meningiomas demonstrated 3D chords and whorls. Low-grade meningothelial nuclei showed streaming and displayed regularity in size, shape, and distribution. Oligodendrogliomas showed regular round nuclei and a variably staining background. Glioblastomas showed high degrees of nuclear pleomorphism and disarray. Mitoses, vascular proliferation, and necrosis were evident. We demonstrate the utility of a 3D-printed, flexible probe microscope for high-resolution microscopic imaging with increased architectural detail. Enhanced invivo imaging using this device may improve our ability to detect and decrease microscopic tumor burden at excision margins during neurosurgical procedures.

Diagnostic utility of cell blocks and an immunomarker panel in the cytological evaluation of serous effusions

Introduction: Cell blocking (CB) has gained wide acceptance as an integral diagnostic tool in cyto-diagnosis. It allows retrieval and concentration of exfoliated cells in fluid specimens to form paraffin blocks that are interpreted in a biopsy like fashion. The aim of this study was to compare the diagnostic utility of CB to conventional smears (CSs) in the evaluation of serous effusions and to evaluate whether the tested immunomarker panel increases the diagnostic yield of malignancy. Materials and Methods: Serous effusion specimens were processed using both techniques; CSs and CB. An immunomarker panel (Ber-EP4, EMA E29, D2-40 and desmin) was then applied on CB sections. Results: CB enhanced the cellularity; it increased the diagnostic yield by 27.4% over CSs. Both nuclear and cytoplasmic details were better demonstrated in CBs. CBs were superior in the preservation of architectural patterns as acini, papillae and sheets. The diagnosis of CBs concurred with that of CSs in 79% of cases, however the diagnostic yield of malignancy increased by 9.6% using CBs. Adding the immunomarker panel totally eliminated the suspicious category and further increased the diagnostic yield for malignancy by 9.7% over CBs and by 19.3% over CSs. Furthermore, categorization of malignant cases into primary malignant mesothelioma and metastatic carcinoma was feasible after thorough interpretation of the combined immunoprofile. Conclusion: CB represents a cost-effective adjunct to CSs in the evaluation of serous effusions. It increases the cellular yield, provides superior cytomorphology, architectural preservation, and supplies material for ancillary studies. Keywords: Serous effusions, Cell blocks, Immunohistochemistry, Marker panel.

The performance of digital microscopy for primary diagnosis in human pathology: a systematic review.

Validation studies of whole slide imaging (WSI) systems produce evidence regarding digital microscopy (DM). This systematic review aimed to provide information about the performance of WSI devices by evaluating intraobserver agreement reported in previously published studies as the best evidence to elucidate whether DM is reliable for primary diagnostic purposes. In addition, this review delineates the reasons for the occurrence of discordant diagnoses. Scopus, MEDLINE/PubMed, and Embase were searched electronically. A total of 13 articles were included. The total sample of 2145 had a majority of 695 (32.4%) cases from dermatopathology, followed by 200 (9.3%) cases from gastrointestinal pathology. Intraobserver agreements showed an excellent concordance, with values ranging from 87% to 98.3% (κ coefficient range 0.8-0.98). Ten studies (77%) reported a total of 128 disagreements. The remaining three studies (23%) did not report the exact number and nature of disagreements. Borderline/challenging cases were the most frequently reported reason for disagreements (53.8%). Six authors reported limitations of the equipment and/or limited image resolution as reasons for the discordant diagnoses. Within these articles, the reported pitfalls were as follows: difficulties in the identification of eosinophilic granular bodies in brain biopsies; eosinophils and nucleated red blood cells; and mitotic figures, nuclear details, and chromatin patterns in neuropathology specimens. The lack of image clarity was reported to be associated with difficulties in the identification of microorganisms (e.g., Candida albicans, Helicobacter pylori, and Giardia lamblia). However, authors stated that the intraobserver variances do not derive from technical limitations of WSI. A lack of clinical information was reported by four authors as a source for disagreements. Two studies (15.4%) reported poor quality of the biopsies, specifically small size of the biopsy material or inadequate routine laboratory processes as reasons for disagreements. One author (7.7%) indicated the lack of immunohistochemistry and special stains as a source for discordance. Furthermore, nine studies (69.2%) did not consider the performance of the digital method-limitations of the equipment, insufficient magnification/limited image resolution-as reasons for disagreements. To summarize the pitfalls of digital pathology practice and better address the root cause of the diagnostic discordance, we suggest a Categorization for Digital Pathology Discrepancies to be used in further validations studies. Among 99 discordances, only 37 (37.3%) had preferred diagnosis rendered by means of WSI. The risk of bias and applicability concerns were judged with the QUADAS-2. Two studies (15.4%) presented an unclear risk of bias in the sample selection domain and 2 (15.4%) presented a high risk of bias in the index test domain. Regarding applicability, all studies included were classified as a low concern in all domains. The included studies were optimally designed to validate WSI for general clinical use, providing evidence with confidence. In general, this systematic review showed a high concordance between diagnoses achieved by using WSI and conventional light microscope (CLM), summarizes difficulties related to specific findings of certain areas of pathology-including dermatopathology, pediatric pathology, neuropathology, and gastrointestinal pathology-and demonstrated that WSI can be used to render primary diagnoses in several subspecialties of human pathology.

The Diagnostic Utility of Cell Block Preparation with Conventional Cytological SmearsA Cross Sectional Study

Background: Cytological examination of serous fluids aspirated is a simple and relatively non-invasive technique to diagnose whether the effusion is malignant or benign. Cell block preparation along with conventional smear increases the sensitivity of detecting malignancies, and also has the ability to reduce false-positive interpretations. Methods: A total 68 samples of body fluid (pleural and ascitic) specimens were examined for conventional cytological smear (CS) and cell block method (CB) over a period of one year. Out of 68 fluids, 40 were pleural fluid and 28 were ascitic fluid. Each fluid specimen was examined by conventional smear technique as well as cell block technique. The morphological details, cellularity, architecture, nuclear and cytoplasmic details were studied in both CS and CB techniques. Result: A total 82.35% smears had adequate material; while of the total cell blocks, 75% cell blocks had adequate material. A total of 11.76% cases were malignant on smears,5.88% were suspicious of malignancy, 64.7% were benign/non-neoplastic lesions. A total 13.2% cases were malignant on cell block, 1.47% were suspicious of malignancy, 60.29% were benign/non-neoplastic lesions. Sensitivity, positive and negative predictive value and accuracy of cell block technique were greater than that of FNAC smears Conclusion: For the final cytodiagnosis of body fluid, there is statistically significant difference between the two techniques. Cell blocks prepared from the residual fluid specimen can be useful for more definitive diagnosis, with advantage of IHC and special stains where required.

Comparison of Conventional Cytology, Liquid-Based Cytology, and Cell Block in the Evaluation of Peritoneal Fluid in Gynecology Malignancies

Background: Peritoneal washing cytology has become an accepted method in evaluating gynecology malignancies. Objective: The aim of this work was to compare the conventional cytology, liquid-based cytology (LBC), and cell block in the evaluation of peritoneal fluid. Materials and Methods: A total of 48 cases with ovarian and uterine malignancy were included in this study. The age of the patients varied from 15 to 71 years. All the 48 cases were subjected to conventional smear, LBC, and cell block preparation. Results: The results of LBC and conventional methods were in line with all cytological criteria, except for the background (p = 0.045), but no significant difference was found regarding adequacy (p = 0.12), cellularity (p = 0.13), cell architecture (p = 0.751), nuclear details (p = 0.96), and cytoplasmic details (p = 0.32). The kappa correlation between conventional cytology and LBC, conventional cytology and cell block, and LBC and cell block was 0.769, 0.791, and 0.945, respectively. The most prevalent malignancy which led to peritoneal fluid involvement was papillary serous carcinoma of the ovary. Conclusion: Compared to conventional cytology, the liquid-based method had no significant superiority in the evaluation of the peritoneal fluid method, but the combined usage of smears and cell block improved the diagnostic accuracy of the peritoneal washing samples in different gynecological malignancies.

Comparison between cytospin and liquid-based cytology in cerebrospinal fluid diagnosis of neoplastic diseases: A single institution experience.

The current tools available for detecting malignant neoplasms in the cerebrospinal fluid (CSF) are neurological examination, followed by neuroimaging, cytology and molecular techniques. To highlight the role of cytology the diagnosis of metastatic tumours in CSF samples, we present our experience using cytospin and ThinPrep liquid-based cytology. A retrospective analysis was conducted using the pathological records of 8181 cytological specimens of CSF, which were diagnosed over a 17-year period. Between 2000 and 2014, a total of 6994 CSF samples were processed using cytospin method and 1187 specimens were examined using ThinPrep method in the period between 2015 and 2017. The most frequent metastatic neoplasm of the first period was non-Hodgkin lymphoma; in the second period the commonest malignancy found was brain tumour (glioblastoma and medulloblastoma). The samples processed by cytospin revealed cytolysis and haemorrhage, while the cases processed by ThinPrep had a clear background. Ten false-positive cases belonging to the suspicious category were processed by cytospin, while there was only one false positive case in the group processed by ThinPrep. The positive predictive value was 95% in cytospin and 100% in Thin Prep with comparable sensitivity, specificity, diagnostic accuracy and negative predictive values. CSF cytology is a reliable technique for identifying malignancy in CSF. ThinPrep technology can be applied with good results in terms of clear background, cell enrichment, better nuclear details and high cellularity per slide.

Liquid Paraffin as a Rehydrant for Air Dried Buccal Smear

Aim: To assess the efficacy of liquid paraffin as a rehydrant for air dried buccal smear based on the nuclear and cytoplasmic details. Background: Buccal smear is useful for diagnosing Malignancy, Fungal infection, Viral infection and Vesicullobullous dermatoses. The routine practice is to wetfixthe smear and send them to the laboratory for staining and evaluation by a cytopathologist. But drying of smears is inevitable, especially when the aspirate is less and when there is improper fixation. This can cause drying artifacts. An alternative method for overcoming this problem is intentional air drying followed by rehydration. Many rehydrants have been experimented upon. The most common rehydrant being saline. The present study is done to check the efficacy of liquid paraffin as a rehydrant. Material and methods: 2 sets of 20 buccal smears were collected from the patients. One set air dried for 24 hours and the other wet fixed. Conventional pap staining was then carried out. The slides were examined by two observers for preservation of Nuclear and cytoplasmic details based on the semi quantitative scoring system. Results: Excellent nuclear details were seen in 45% of air dried smear compared to 25%in routine wet fixation. The cytoplasmic details of air dried smear and routine wet fixed smear was the same. Conclusion: Liquid paraffin as a rehydrant has shown promising results and the nuclear details were found to marginally better than in routine wet fixed smear. Air dried buccal smears can be used in routine practice especially in rural areas where there is limited access to laboratories and for mass screening.

Comparison of Liquid-based and Conventional Cytology in Diagnosis of Breast Mass.

Background:Among all diagnostic techniques for breast lesions, fine-needle aspiration (FNA) is the simplest, most reliable and cheapest one.Aim:To compare liquid-based and conventional cytology in diagnosis of breast mass.Materials and Methods:About 101 patients with breast mass were enrolled. The aspirated materials were divided into two parts. One was poured into a liquid medium and the other part was directly spread on clean glass slides. Conventional and liquid-based preparations were compared using several criteria including adequacy (presence of the epithelial cluster or myoepithelial cells), overall cellularity, presence of single epithelial cells, presence of myoepithelial cells, epithelial architecture, nuclear detail, nuclear atypia and inflammatory/proteinaceous background and final diagnosis.Results:Among 101 cases, 85 (84.1%) were malignant and 16 (15.9%) were benign. Conventional and liquid-based cytology were similar according to adequacy (P = 0.65), cellularity (P = 0.13), epithelial architecture (P = 0.15), presence of myoepithelial cells (P = 0.61), nuclear detail (P = 0.57) and nuclear atypia (P = 0.44), while there were a significant difference between the two methods according to the presence of single epithelial cells (P < 0.001) and background (P < 0.001).Conclusion:Liquid-based cytology of breast specimens is an accurate diagnostic tool with high diagnostic yield in benign and malignant lesions.

Liquid-based cytology versus conventional cytology in fine-needle aspiration cytology of thyroid, salivary gland, and lymph node swellings: a comparative study

Purpose To compare the cytomorphologic features of liquid-based cytology (LBC) and conventional preparation (CP) smears in fine-needle aspiration of thyroid, lymph node, and salivary glands. Patients and methods The study included 110 patients experiencing thyroid, salivary gland, and lymph node swellings, who presented to the Surgical Oncology Outpatient Clinic and referred to Cytology Unit of the Pathology Department of the National Cancer Institute, Cairo University, for fine-needle aspiration cytology, from the period January 2017 till July 2018. For each fine-needle aspiration cytology, three passes were performed: the first two were for conventional cytology, and the third one for LBC, where a semiautomated SurePath technique was used. The comparison between both techniques was done using Wilcoxon signed rank test, regarding the cellularity, cell architecture, monolayering, nuclear details, cytoplasmic details, informative background, and bloody background. Results CP was significantly better regarding the cellularity, cellular architecture, and informative background of thyroid lesions, whereas LBC was better when the absence of bloody background was considered. On the contrary, no significant difference was found between both preparations regarding monolayer formation and nuclear and cytoplasmic details. Conclusion CP is superior to LBC in preservation of cellular pattern, gives better cellular yield, eliminates hemorrhagic background, and maintains the stroma and background, which are important for diagnosis. As such stroma is lacking in LBC, it should be interpreted with support of CP in certain lesions.

Comparative Analysis of Liquid based and Conventional Cytology Smears in Fine Needle Aspirates from Breast Lesions.

Context:Liquid-based cytology (LBC) is being extensively used for the evaluation of both gynecological and non-gynecological specimens. Suspension of cells in monolayer makes better morphological assessment possible. Along with this, inherent morphological changes such as altered, reduced, or lost background material, fragmented cell clusters, smaller cell size, nucleolar prominence, etc., need to be considered.Aim:Present study was aimed at comparative evaluation of utility of LBC versus conventional smear (CS) in assessing breast lesions and whether it can be used as an alternative to conventional preparation.Settings and Design:Present study was a prospective study in which 75 cases of breast fine-needle aspiration cytology from patients with palpable breast lumps constituted the study group.Material and Methods:The first pass was used for CS and LBC; a second pass was given. The representative CS and LBC smears were compared using several criteria.Statistics:Each feature was scored individually and evaluated statistically using Wilcoxon's signed rank test on the SPSS program.Results:A statistically significant difference was found in informative background and background blood-debris, whereas the difference was not statistically significant in other features such as cellularity, cytoarchitectural pattern, presence of monolayer, and nuclear and cytoplasmic details.Conclusion:LBC is a promising technique in the field of cytology. It has the potential to decrease the number of slides screened per case and decrease the turn-around-time.

Cocktail of Periodic Acid-Schiff and Papanicolaou: Novel staining technique for the identification of leukemic eosinophils - A pilot study.

Background:Tissue eosinophilia may be caused due to reactive, neoplastic or idiopathic reasons. Reactive eosinophils in allergic and inflammatory conditions are transient and recruited from the circulation in response to various stimuli, whereas neoplastic eosinophils of leukemias and hematological malignancies are involved in the pathogenesis of the disease. The differentiation of reactive from neoplastic eosinophils has a serious implication on the treatment and prognosis of diseases. However, both these types of eosinophils display variation in morphology and staining characteristics in routine histopathology leading to a diagnostic dilemma.Aim:The aim of this study is to evaluate the efficacy of special stains for the demonstration of eosinophils in normal/reactive lesions and leukemias.Methodology:A retrospective study comprising twenty histologically diagnosed cases each of reactive oral lesions and leukemias were obtained from institutional archives. These tissue sections were subjected to staining with routine and special stains – Carbol chromotrope, Congo red, Leishman's stain, Periodic Acid–Schiff-papanicolaou (PAS-PAP) and PAS. Statistical analysis was performed using Pearson's Chi-square test to compare the various parameters in the evaluation of the staining efficacy.Results:Carbol chromotrope and Congo red staining showed increased staining efficacy in normal/reactive eosinophils while PAS-PAP followed by PAS and Leishman's stain showed enhanced features such as homogenicity, specificity, increased staining intensity, enhanced nuclear and cytoplasmic details in leukemic eosinophils.Conclusion:Combined PAS-PAP is a novel and cost-effective staining technique in differentiating reactive and leukemic eosinophils. It is significant in recognizing leukemic eosinophils of routine biopsies and alerts the clinician to rule out any underlying malignancies.

Pathomorphology of malignant melanoma in a canine spleen

A case of malignant melanoma was recorded in a four year old female Rottweiler. Grossly, four subcapsular smooth nodules of varying sizes were observed on the parietal surface and one smooth nodule on the visceral surface. On section, the nodule was soft, dirty grey-black, circumscribed and appeared as two distinct areas separated by parenchyma. Cytology revealed oval to polygonal cells containing black granular cytoplasmic pigments, obscuring the nuclear details. Anisocytosis, anisokaryosis, variable nuclear:cytoplasmic ratio, prominent nuclei were seen. Histopathologically, the neoplastic cells were round to spindle to polyhedral in shape with severe/deep and brown black pigmented cytoplasm. On special staining with Fontana, the granules of neoplastic cells stained black and nuclei stained pink. The tissue was then subjected to proliferation marker Ki-67 immunohistochemistry, where more than 75% of neoplastic cells showed strongly positive immunolabelling.

Efficacy of acridine orange and papanicolaou stains in sex determination using barr bodies in buccal smears: A comparative study

Objective: Sex determination can be done by buccal epithelial cells in saliva traces found at a crime scene by examining the presence of Barr bodies in the nucleus of the epithelial cells. The present study aims to assess the efficacy of sex determination using Acridine Orange (AO) and Papanicolaou (PAP) stains for the detection of Barr bodies in buccal mucosal scrapings. Materials and Methods: Buccal scrapings from 120 healthy individuals (60 males and 60 females) were collected and were stained with AO and PAP stains. The analysis of fifty epithelial cells in each sample was done for the identification of Barr bodies. The presence of Barr bodies ≤ 5% in the sample was recorded as male and those with > 5% was recorded as female. Both stains were evaluated for percentage accuracy in determining sex. Results: In AO-stained slides, the percentage of Barr bodies ranged from 4 to 31 in females and from 0 to 9 in males, whereas with PAP the ranges recorded were 3–21 in females and 0–6 in males (P Conclusion: Using Barr bodies in the buccal cells, providing up to 95%–98% accuracy, made it a considerable aid for sex determination. AO stain proved better than PAP stain for visualizing nuclear details with its remarkably shorter staining time and confirmed superior sex estimation efficiency compared to PAP stain.

Comparison of Ultrafast Papanicolaou stain with standard Papanicolaou stain for cervical smear

Background: After the first evolution of Papanicolaou smear, it has been successfully used to screen cervical cancer. With modifications in the staining method of Pap stain, the lesser staining time and unequivocal cell morphology has been achieved by Ultrafast Papanicolaou stain. The aim of this study is to compare the UFP stain and SP stain in cervical pap smears on the basis of background, cell morphology, nuclear details and overall staining.Materials and methods: This is a prospective hospital based study conducted in a tertiary hospital, Nepal Medical College over a period of six months (January 2017 to June 2017). Cervical pap smears received in the department of Pathology is subjected to Standard papanicolaou (SP) stain and Ultrafast papanicolaou (UFP) stain for comparison.Results: Total of 368 cervical pap smears were subjected to SP stain and UFP stain. Mean age of the patient was 36.57+-10.06 years. The quality of UFP stain was assessed by considering the background, cell morphology, nuclear details and overall staining. Difference in regards to the interpretation and background was not significant when SP stain was compared to UFP stain. However, cell morphology, nuclear details and overall staining were far better in smears stained with UFP stain.Conclusion: Pap smear is the simple and effective screening tool for cervical lesions. With modifications in staining method and introduction of UFP stain, the result is lesser staining time and excellent morphological quality. Hence, UFP stain has become the choice of stain and can be used on a regular basis.

Optimizing optical coherence tomography and histopathology correlation in retinal imaging

ObjectiveTo develop a methodology to correlate optical coherence tomography (OCT) images and histopathological sections from the same eye. Part 1: To determine the best fixative for optimal OCT and histopathological analysis in post-mortem eyes. Part 2: A protocol is proposed to correlate histopathological features and OCT scans from the same post-mortem eyes. DesignExperimental study. ParticipantsPart 1: Twenty-three rabbit eyes and 14 post-mortem human eyes. Part 2: Nineteen post-mortem human eyes. MethodsPart 1: Six different fixatives were tested, and specimens were evaluated on 4 criteria: globe shape, structure opacification, retinal detachment, and nuclear details. Part 2: Based on the findings from Part 1, fixed human eyes were imaged using OCT. Orientation-controlled histopathological processing was performed to obtain serial tissue sections from paraffin embedded tissue, which were matched to corresponding OCT images. ResultsPart 1: Of the 6 fixatives, 2% glutaraldehyde and Davidson's solution met the proposed criteria in rabbit eyes. Of these, glutaraldehyde showed similar results in human eyes and was selected for Part 2. Part 2: Using anatomical landmarks, cross-sectional histopathological sections in the same orientation as the OCT images were correlated to their corresponding OCT images. Retinal lesions such as a macular hole, an epiretinal membrane, and the presence of drusen were easily correlated, proving the reliability of our methodology. Moreover, the photoreceptor's inner/outer junction was correlated to a hyperreflective band on OCT. ConclusionsA standardized protocol was developed to correlate OCT images and histopathological findings by generating serial cross-sections of the retina, which can be used to better understand otherwise ambiguous OCT findings.

Liquid dish wash solution – can it be an alternative in future for the expensive and hazardous xylene in hematoxylin and eosin staining of paraffin sections

Introduction: Xylene is the most commonly used clearing agent as it has excellent deparaffinising and clearing properties. Commercial xylene is a clear, colourless liquid that has a sweet and aromatic odour. However, xylene is not only expensive but also has detrimental effect on human health such as hepatitis, chemical pneumonitis, depression, anaemia, etc. Hence in this study we are replacing xylene with commercially available liquid dish wash solution (LDW) which is not only cheap but also non biohazardous. Aims: To assess the efficacy of LDW with xylene as a deparaffinising and clearing agent in routine hematoxylin and eosin staining (H&E). Materials and Methods: Blocks from 20 consecutively submitted samples at RL Jalappa Hospital between Jan – June 2015 each of breast, Cervix, Lymphnode, Fat, Salivary Gland, Muscle and Alimentary Tract was taken and two sections of 4-micron thickness were cut and deparaffinised using Liquid Dish Wash and Xylene respectively followed by H & E Staining. The 280 sections of the 140 blocks were evaluated and scored by two pathologists independently for nuclear staining, cytoplasmic staining, uniformity, clarity and crispness. Results: In the samples evaluated LDW seems to be on par with xylene cleared sections in terms of nuclear and cytoplasmic detail, clarity and crispness. However Xylene cleared sections showed slightly better uniform staining. However both sections were equally good for reporting. Conclusion: Liquid dish wash solution is an excellent alternative to xylene in H&E Staining. However more extensive studies need to be done on other types of tissues for validation

MP19-18 THE USE OF FORMALIN AS A FIXATIVE FOR TESTICULAR BIOPSIES

You have accessJournal of UrologyInfertility: Epidemiology & Evaluation II1 Apr 2018MP19-18 THE USE OF FORMALIN AS A FIXATIVE FOR TESTICULAR BIOPSIES James Ellenburg, Joseph Drwiega, Anna Posey, Matthew Goldberg, Jennifer Gordetsky, and Peter Kolettis James EllenburgJames Ellenburg More articles by this author , Joseph DrwiegaJoseph Drwiega More articles by this author , Anna PoseyAnna Posey More articles by this author , Matthew GoldbergMatthew Goldberg More articles by this author , Jennifer GordetskyJennifer Gordetsky More articles by this author , and Peter KolettisPeter Kolettis More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2018.02.667AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Testicular biopsy (TB) is important in the management of the infertile man. Bouin solution (BS) has traditionally been the recommended fixative for TB specimens. While buffered formalin (BF) is the most common fixative used in pathology, its use for a testicular fixative has been discouraged due to concern that it may cause histologic artifact that could hamper assessment of spermatogenesis. We evaluated TB fixed in BF and those fixed in BS to assess any difference in histology quality. METHODS 103 patients who underwent TB from January 2000 through December 2016 were identified. Representative slides from 12 patients, 6 fixed with BF and 6 fixed with BS, were evaluated by a genitourinary (GU) pathologist blinded to the fixative used. Nuclear, cytoplasmic, and basement membrane (NM, CM, BM) detail, and nuclear and cytoplasmic granularity (NG, CG), were graded as crisp (high quality) or blurred (low quality). In an attempt to minimize aging artifact in specimens, an additional two pairs of rat testicles were harvested. Each testis was sectioned in half; one half was fixed in BF and one half in BS to yield a total of 8 slides. These were evaluated by a GU pathologist blinded to the fixative used for each case using criteria above. RESULTS Of 71 patients, 56 (78.8%) specimens were fixed in BF, 8 (11.3%) in Zinc formalin, and 7 (9.86%) in BS. 22 slides from 6 specimens fixed in BS and 12 slides from 6 specimens fixed in BF were graded based on above quality metrics. NM detail was crisp in all slides fixed in BF (12/12) and in 10 of 22 slides fixed in BS. NG was crisp in all slides fixed in BF (12/12) and in BS (22/22). CG was crisp in 10 of 12 BF slides and in 16 of 22 BS slides. CM detail was crisp in all BF fixed slides and in 20 of 22 BS fixed slides. BM detail was crisp in all BF slides (12/12) and in 12 of 22 BS slides. For the rat testes, the BF fixed tissue was described as crisp for each metric in all four slides (4/4). The BS fixed tissue had crisp CM detail (4/4), crisp CG (4/4), and BM detail (4/4). However, the NM detail was blurred in half of the tissue (2/4), and the NG was blurred in 3 of 4 slides. CONCLUSIONS Slides from BF fixed specimens were found to be high quality in the evaluated metrics. CG was the only metric that was found to be low quality in one specimen fixed in BF. BS slides demonstrated more variability in quality across all metrics, with each specimen having at least one metric that was found to be blurred, or low quality. These findings were demonstrated in the rat testes as well. We conclude that spermatogenesis and testicular histology can be adequately evaluated with formalin fixation. © 2018FiguresReferencesRelatedDetails Volume 199Issue 4SApril 2018Page: e250-e251 Advertisement Copyright & Permissions© 2018MetricsAuthor Information James Ellenburg More articles by this author Joseph Drwiega More articles by this author Anna Posey More articles by this author Matthew Goldberg More articles by this author Jennifer Gordetsky More articles by this author Peter Kolettis More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Rehydration before wet fixation in conventional body fluid cytology - An 18-year experience.

In conventional cytology, preparation of a specimen by wet fixation for Papanicolaou stain is potentially subject to dry effect or cell loss which may make cytologic interpretation difficult or even impossible. We have been routinely making an additional smear for rehydration with normal saline (rehydration method) before wet fixation to overcome the above shortcomings. We reviewed malignant pleural effusion and ascites 15 cases each in our cytology laboratory over the past 1year. Four slides of each specimen were made. Two were air-dried for Liu's stain (a Romanowsky stain) and the other two were wet-fixed for Papanicolaou stain. The air-dried smears were also served as retained cellularity control. One of the two wet-fixed smears was processed as a control of preservation of nuclear detail whereas the other one stayed air-dried for 10minutes and then covered with normal saline (rehydration method) for 80seconds before wet fixation. There was minor cell loss (P=.032). The cells appeared larger with good preservation of nuclear detail (P<.0001 by two-sided Wilcoxon rank sum test) but no red blood cells retained on the slide after rehydration. The rehydration method can effectively minimise cell loss, enlarge and preserve the cytological features of malignant cells with haemolysis. This method is simple, practical and good for cytological screening for tumour cells and interpretation especially in a bloody smear. We recommend that the rehydration method be part of traditional cytopreparatory work of wet fixation for Papanicolaou stain in conventional body fluid cytology.

Application of the Mesolens for subcellular resolution imaging of intact larval and whole adult Drosophila.

SummaryIn a previous paper, we showed a new giant lens called the Mesolens and presented performance data and images from whole fixed and intact fluorescently‐stained 12.5‐day old mouse embryos. Here, we show that using the Mesolens we can image an entire Drosophila larva or adult fly in confocal epifluorescence and show subcellular detail in all tissues. By taking several hundreds of optical sections through the entire volume of the specimen, we show cells and nuclear details within the gut, brain, salivary glands and reproductive system that normally require dissection for study. Organs are imaged in situ in correct 3D arrangement. Imaginal discs are imaged in mature larvae and it proved possible to image pachytene chromosomes in cells within ovarian follicles in intact female flies. Methods for fixing, staining and clearing are given.