Objective
Buffer‐based cell media (BBCM) are a valuable tool in the post‐collection processing of cytology samples, though with poorly defined effects on cell properties. In this study, time‐related ...changes in cell morphology and biomarker immunoreactivity were evaluated for cells stored at room temperature in a BBCM prepared with bovine serum albumin (BSA) and ethylene diamine tetraacetic acid (EDTA).
Methods
Cytospins were prepared at five consecutive 24‐hour intervals (0, 24, 48, 72, 96) from three human cell lines (MCF7, SK‐MEL‐28, FaDu) suspended and stored in BBCM. Preservation of cell morphology was evaluated on Papanicolaou‐stained cytospins from the percentages of apoptotic cells. Preservation of immunoreactivity was evaluated for cytokeratins, oestrogen receptors, Ki67, and melanoma markers from the percentages of cells positive for the corresponding immunocytochemical reactions.
Results
Cell morphology was well preserved for the majority of cells of the three lines stored for 24 and 48 hours (93%, 97%, 98% and 62%, 81%, 88%, respectively), while the majority of cells were apoptotic after 72 and 96 hours (70%, 47%, 39% and 77%, 70%, 59%, respectively). The immunoreactivity of cytokeratins remained unchanged during the entire 96 hours, while that of melanoma markers (S100, HMB45, Melan‐A) decreased by 27%, 2%, and 3%, respectively. The immunoreactivity of oestrogen receptors and Ki67 decreased by 29% and 17% after the first 24 hours, and was completely lost after 96 hours.
Conclusions
A BBCM with the addition of BSA and EDTA facilitates good preservation of cell morphology and immunoreactivity of biomarkers for up to 48 hours at room temperature.
Buffer‐based media are a valuable tool in post‐collection processing of cytology samples, though with poorly defined effects on cell properties This study confirmed that our in‐house prepared buffer‐based medium with addition of BSA and EDTA facilitates good preservation of cell morphology as well as immunoreactivity of biomarkers' for up to 48 hours at room temperature.
Endoscopic ultrasound-guided fine needle aspiration biopsy (EUS FNAB) is a well established diagnostic method in adult patients, but is rarely used in the paediatric population. The Clinical ...Department of Gastroenterology at the University Clinical Centre Ljubljana and the Department of Cytopathology at the Institute of Pathology, Faculty of Medicine, University of Ljubljana, Slovenia, have been closely collaborating on EUS FNAB since the introduction in 2010. The aim of the study was to review the cases of EUS FNAB of pancreatic neoplasms in children.
In the digital archive of the Institute of Pathology (IP), Faculty of Medicine (FM), University of Ljubljana (UL), we found 6 cases of EUS FNAB in children, 3 had EUS FNAB of the pancreas, 2 of whom had a cytopathologic diagnosis of a tumour. In the first case, the lesion was ultrasonographically solid, and the cell sample contained branching papillary structures surrounded by aggregates of small cells with nuclear grooves. In the second case, the lesion was ultrasonographically cystic, and predominantly necrosis was seen, with only single preserved cells. Positive nuclear reaction for β-catenin was found in both cases by immunohistochemical staining.
In both cases, the cytopathological diagnosis of solid pseudopapillary neoplasm of the pancreas was made, the cases represent the totality of paediatric cases of pancreatic neoplasms from the Children's Hospital Ljubljana since 2010. There were no adverse events during and after EUS FNAB. A histopathological examination of the tumour resection specimens confirmed the cytopathological diagnosis.
Our experience indicates that EUS FNAB is a safe and effective method for diagnosing pancreatic neoplasms in the pediatric population, as supported by the findings in the literature.
Background
There are numerous methods and procedures described for the preparation of cell blocks (CBs) from cytological samples. The objective of this study was to determine current practices and ...issues with CBs in European laboratories.
Methods
A link to an online survey, with 11 questions about CB practices, was distributed to cytology laboratories via participants of United Kingdom National External Quality Assurance Service for Cellular Pathology Techniques and national representatives in the European Federation of Cytology Societies.
Results
A total of 402 laboratories responded completely (337/402, 84%) or partially (65/402, 16%) to the survey by February 4, 2022. The most common CB practice is embedding cell pellets using plasma and thrombin (23.3%), agar (17.1%), Shandon/Epredia Cytoblock (11.4%), HistoGel (7.9%), and Cellient (3.5%). Other methods such as CytoFoam, albumin, gelatin, Cytomatrix, and collodion bags are rarely used (1.0%, 0.7%, 0.7%, 0.3%, and 0.2%, respectively). CBs are also prepared from naturally occurring clots or tissue fragments (29.5%) and cells scraped from unstained or prestained smears (4.4%). The most frequent issues with the CBs in a daily cytology practice are low cellularity (248/402, 62%) and dispersed cells (89/402, 22%), regardless of the CBs preparation method or how the samples for embedding were selected.
Conclusions
There is a great variability in CB practices in European laboratories with low cellular CBs as the main issue. Additional studies are mandatory to evaluate and improve performance and cellular yield of CBs.
There is great variability in cell block (CBs) practices in European laboratories with low cellular CBs as the main issue. Additional studies are needed in order to evaluate and improve performance and cellular yield of CBs.
Introduction
The aim of this retrospective study was to evaluate the preservation of biomarkers immunoreactivity on cytospins protected with polyethylene glycol (PEG).
Methods
In two independent ...cytopathology laboratories, immunocytochemical reactions were retrospectively evaluated on methanol‐fixed and PEG‐protected cytospins stored at room temperature (RT) for different time periods and compared with immunocytochemical reactions on corresponding baseline methanol‐fixed cytospins. Semi‐quantitatively assessed immunoreactivity, using scores from 0 to 3, was considered reduced if two sequential scores were lowered by at least one point.
Results
Immunocytochemical reactions for 40 biomarkers with membrane (10), cytoplasmic (22) and nuclear (8) localisation were performed on 921 slides prepared from 183 cytological samples. For the majority of biomarkers (29/37, 78%), immunoreactivity on PEG‐protected cytospins stored at RT remained unchanged in the first 12 months. Immunoreactivity for GFAP, p40 and hepatocyte antigen was monitored and remained unchanged for 1, 8 and 7 months, respectively. Partial or complete loss of immunoreactivity on PEG‐protected cytospins stored for less than 12 months was found on a single sample out of the total evaluated for CD3 (1/7), CD30 (1/4), CD45 (1/10), CK5/6 (1/7), MelanA (1/7) and vimentin (1/7), while more frequent changes of immunoreactivity were found for Ki67 (4/7) and p63 (2/7).
Conclusion
Immunoreactivity on cytospins protected with PEG and stored at RT is well‐preserved for at least 12 months for the majority of biomarkers.
Cell morphology and immunoreactivity of biomarkers is well preserved for at least 12 months on methanol‐fixed cytospins protected with polyethylene glycol and stored at room temperature. Appropriate cytology control slides can be prepared for immunocytochemistry on cytology samples
Erythroblastic sarcoma (ES) is an extremely rare extramedullary solid tumor of immature erythroid cells, with around a dozen human cases to our knowledge presented so far in the english literature, ...none of the cases presenting as pleural effusion. Here we describe a case of ES in a thoracic wall diagnosed by cytological examination of pleural effusion with histologic correlation in a 53‐year‐old patient with a 10‐month history of myelodysplastic syndrome/myeloproliferative neoplasm with grade III reticulin myelofibrosis.
Light microscopic evaluation of cell morphology in preparations from urine or bladder washing containing exfoliated cells is a standard and primary method for the detection of bladder cancer and also ...malignancy from other parts of the urinary tract. The cytopathologic examination is a valuable method to detect an early recurrence of malignancy or new primary carcinoma during the follow-up of patients after the treatment of bladder cancer.
Characteristic cellular and nuclear signs of malignancy indicate invasive or in situ urothelial carcinoma or high-grade papillary urothelial carcinoma. However, low sensitivity of the method reflects the unreliable cytopathologic diagnosis of low-grade urothelial neoplasms as cellular and nuclear signs of malignancy in these neoplasms are poorly manifested. Many different markers were developed to improve the diagnosis of bladder carcinoma on urinary samples. UroVysion™ test is among the newest and most promising tests. By the method of in situ hybridization one can detect specific cytogenetic changes of urothelial carcinoma.
Thyroid-like follicular carcinoma of the kidney (TLFC), a rare neoplasm with low malignant potential, is histologically similar to primary thyroid follicular carcinoma, but characteristically lacks ...thyroid immunohistochemical markers. We report a case of 34-year old patient with nephrolithiasis. Ultrasound revealed hepatorenal cysts consistent with adult type polycystic kidney disease (ATPKD) and a cytologically confirmed left kidney tumor. Nephrectomy specimen contained sharply demarcated lesion of unusual morphology. Tubular and cystic structures lined by mostly cuboidal cells and filled with amorphous eosinophillic material, reminiscent of follicular carcinoma of the thyroid gland, were diagnostic for TLFC. Thyroid markers were negative. To our knowledge this is the first report of TFLC associated to ATPKD. Brief review of previously published TFLCs, possible relationship between entities and differential diagnosis are discussed.
The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/8067946569612694.
Background
Different cytology preparations can be used for molecular diagnostics, however the influence of pre‐analytical and analytical steps on the results are not yet well defined. We aimed to ...determine optimal steps for efficient extraction of DNA and RNA from fresh cells for molecular diagnostics.
Methods
MCF7 and FaDu human cell lines, were used as a model to determine fresh cells storage conditions (temperature: 25°C, 4°C, −20°C, −80°C; duration: 0 h, 4 h, 12 h, 24 h, 48 h) and optimal nucleic acids extraction method. Besides, the minimal number of total cells and minimal percentage of mutated cells needed for successful extraction of nucleic acids and subsequent determination of present mutation were evaluated.
Results
Extraction of nucleic acids using spin columns yielded the highest quantity and quality of nucleic acids. Isolation of nucleic acids was feasible in all storage conditions, however higher temperature and longer duration of fresh cells storage were associated with lower quality of isolated nucleic acids and similar quantification cycle of housekeeping genes. Successful molecular testing was feasible with least 104 cells, while specific mutation was detected in as low as 5% of mutated cells.
Conclusions
Our cell line model, mimicking fresh cytology samples, showed that quantity of extracted either DNA or RNA declined with higher temperatures and longer duration of storage but regardless of the storage conditions, we successfully detected both housekeeping genes and mutated gene using qPCR.
Using cell line model to mimic fresh cytology samples, we have determined the influence of storage conditions (time, temperature) on the quality and quantity of nucleic acids for cells suspended in a buffer‐based cell medium. Additionally, we assessed the minimal number of cells and minimal percentage of mutated cells per sample for successful molecular testing.
Pheochromocytomas and sympathetic paragangliomas are rare tumours arising from chromaffin cells, producing catecholamines in various amounts. Fatal hypertensive episodes may occur perioperatively, ...which are preventable by alpha adrenergic receptor blockers. The perioperative mortality rate of diagnosed versus undiagnosed catecholamine‐producing tumours is significant, considering that only a minority of tumours develop metastasis. Herein we describe a case of a primary adrenal pheochromocytoma referred to as a pancreatic tumour, successfully diagnosed by endoscopic ultrasound‐guided fine needle aspiration biopsy, with a distinct morphology (prominent nuclear anisonucleosis, intranuclear pseudoinclusions, and multinucleation) and immunohistochemical signature.
A case of a primary pheochromocytoma referred to as a pancreatic tumour, successfully diagnosed by endoscopic ultrasound‐guided fine needle aspiration biopsy using GATA3 immunostain.
The aim of this study was to assess signs and symptoms of ocular surface disease (OSD) and the cytomorphological changes of ocular surface in glaucoma patients using preserved antiglaucoma drops.
In ...this cross-sectional study, 109 participants (79 patients with topical medication and 30 untreated controls) completed the Ocular Surface Diseases Index (OSDI) questionnaire and underwent an ophthalmic examination, including Schirmer test, tear film breakup time (TBUT), and fluorescein staining. Conjunctival specimens were collected by impression cytology and analyzed by light microscopy using Nelson's grading scheme (grades 0-3). This classification is based on the nucleus-to-cytoplasm ratios of epithelial cells and the numbers of goblet cells, with grade 2 considered abnormal.
The medication group had significantly shorter TBUT (median interquartile range: 6.0 seconds 5.0-8.0 seconds vs 9.5 seconds 6.0-12.3 seconds; P<0.03), greater fluorescein staining (1.0 0.75-1.25 vs 0 0-0.25; P<0.001), and higher impression cytology grade than the control group (median range: 1.0 1:2 to 1:6 vs 0.6 1:2 to 1:4; P<0.001). The increasing number of drops instilled per day was associated with an increase in fluorescein staining (Spearman's rho r=0.475; P<0.001) and shorter TBUT (r=-0.278; P=0.014). The OSDI did not discriminate between the two groups.
Clinical tests and impression cytology showed ocular surface damage in patients using preserved antiglaucoma medications. However, there was no statistically and clinically significant difference in symptoms measured by OSDI score between the medication and control groups.
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