Cell Block Preparation Research Articles

Endoscopic ultrasound-guided fine needle aspiration cytology of pancreatic adenocarcinomas revisited. A detailed cytological analysis

Background:Early detection of pancreatic adenocarcinomas is essential for improving survival. In this regard, endoscopic ultrasound-guided fine-needle aspiration cytology (EUS-FNAC) has established itself as the method of choice for its ability to target lesions smaller than those which could be targeted by the traditional imaging methods like transabdominal ultrasound. Identifying these tumors correctly on FNA may be challenging because pancreatic adenocarcinomas may show a wide range of morphological features and the presence of contaminants from the gastrointestinal tract may show up as potential pitfalls. This study presents detailed cytomorphological analyses of 59 cases reported as pancreatic adenocarcinomas on smears and cell blocks. The clinical and histopathology follow-up data wherever available have also been presented.Materials and Methods:EUS-FNAC smears and cell blocks from cases reported as pancreatic adenocarcinomas were retrospectively evaluated with individual assessments of a range of features related to cellularity, cellular arrangement, cytoplasmic qualities, and nuclear features. Aspirates from peripancreatic lymph nodes, histopathology sections, and clinical records were reviewed wherever available.Results:Nonneoplastic cells like pancreatic ductal cells and acinar cells, duodenal, and gastric epithelia were detected along with neoplastic cells showing a wide range of variations in different cytomorphological characters. Often, a mixture of features was noted in the same case. Cell block preparations served as useful adjuncts since they made it possible to render unequivocal diagnoses of malignancies in cases where smears were hypocellular.Conclusion:The study creates a useful knowledge base of cytomorphological features of pancreatic adenocarcinomas.

Role of Endobronchial Ultrasound Guided Transbronchial Needle Aspiration with Cellblocks in Diagnosis and Subtyping of Intrathoracic Lesions: Two Year Experience from a Tertiary Care Center.

Background:Endobronchial ultrasound guided transbronchial needle aspiration (EBUS-TBNA) allows precise real-time sampling of intrathoracic lesions and is a minimally invasive, safe, and cost effective technique with high diagnostic yield. The aim of the current study is to evaluate utility of EBUS-TBNA cytology with cellblocks (CBs) in the diagnosis and subtyping of the intrathoracic lesions.Materials and Methods:It was a prospective study conducted from October 2015 to October 2017. We received 233 cases of intrathoracic lesions for EBUS-TBNA, of which CB was made in 217 cases. A flexible endobronchial ultrasound scope was used to sample the lesions.Results:There were 89 females and 128 males amongst the total 217 patients (age range: 14–85 years, mean age: 48.35 years). The smears from EBUS-TBNA contained adequate material in 137/144 [95.1%] non-neoplastic cases and 34/36 [94.4%] of the neoplastic cases, whereas the CBs provided adequate material in 94/144 [65.3%] non-neoplastic cases and 37/40 [92.5%] neoplastic cases. The CB helped subtyping the malignancy in 19 cases and of these immunohistochemistry (IHC) was done on the CB in 15 cases. The biopsy was non-diagnostic in 17/36 cases of granulomatous pathology and 4/18 neoplastic cases diagnosed with EBUS-TBNA.3.Conclusions:EBUS-TBNA provides rapid diagnosis through cytology and the material recovered in the same setting for the CB preparation can be used for immunohistochemical analysis and it may at times provide the diagnosis in cases where the smears are non-diagnostic.

Specimen-specific cell-blocking approaches.

The science of CellBlockistry highlights requirement for proper approach to process different types of cytopathology specimens with critical considerations during preparation of cell-blocks. Common cytopathology specimens which may be subjected for cell-blocking include FNA aspirates in addition to anterior fat pad aspirate, bone marrow aspirate, effusion fluids, and other fluids such as various washings and urine. In addition veterinary sciences and research fields including animal experiments and tissue/cell cultures may also be cell-blocked for improved diagnostic yield and research outcome.

Utility of Fine Needle Aspiration Cytology in the Evaluation of Lymphadenopathy.

IntroductionFine needle aspiration cytology (FNAC) is a quick, effective and relatively inexpensive technique to evaluate the visibly accessible superficial masses. As cervical, axillary and inguinal lymphadenopathies are commonly encountered clinical problems, in this study, we evaluated the utility of FNAC for assessment of lymphadenopathy.MethodsA retrospective observational study was conducted in the Department of Cytopathology, Liaquat National Hospital and Medical College, over the duration of three years. A total of 559 cases were included in the study that underwent FNAC. After palpation, two to three passes were performed with a 22-23 gauge needle along with a plunger for FNAC. The obtained material was spread on three slides that were then stained with hematoxylin and eosin (H & E), Papanicolaou (PAP), and Diff-Quik methods. The remaining material was used for cell block preparation.ResultsThe mean age of the patients was 37.05±18.03 years. In 98.7% of cases, the material was adequate for a satisfactory cytological examination. The most common site of FNAC was the cervical lymph node and tuberculous lymphadenitis (37%) was the most common diagnosis on FNAC, followed by reactive lymphadenitis (27.2%). Reactive lymphadenitis was seen more frequently in the younger age group (<15 years), whereas metastatic carcinoma was more commonly seen in the older age group (>50 years). Tuberculous lymphadenitis was noted more frequently in the middle age group (16-35 years). Moreover, tuberculous lymphadenitis was noted more commonly in cervical lymph nodes, while metastatic carcinoma was more frequently observed in axillary and inguinal lymph node FNACs.ConclusionFNAC is a quick and reliable method to categorize the cause of lymphadenopathy into reactive, inflammatory/infectious, metastatic, and lymphoproliferative, avoiding the necessity of an incisional/trucut biopsy. Moreover, age, gender, and site of involvement are useful predictors of the cause of lymphadenopathy. We noted that in the younger age group, reactive lymphadenitis was more common, whereas tuberculous lymphadenitis and metastatic carcinoma were more frequent in middle and older age groups, respectively. On a similar note, tuberculous lymphadenitis was more frequent in cervical lymph nodes than axillary and inguinal lymph nodes, while metastatic carcinoma was more common in these latter two sites.

Cell blocks in veterinary medicine: A comparison of two methods (cell tube and agar) in 52 effusions from dogs and cats.

Cell blocks are alternative preparations of fluid cytological specimens. They can be used for immunochemical studies as complementary tools or when other techniques (eg, immunocytochemistry, flow cytometry) are not available. We aimed to provide comparative morphologic, immunohistochemical, and technical features of agar-based cell blocks (ACBs) and cell tube blocks (CTBs) from cavitary effusions. Agar-based cell blocks and CTBs were obtained from canine and feline effusions with neoplastic/atypical cells or with packed cell volumes ≥3%. Cellularity, RBC separation, and cellular features were evaluated on digitalized H&E slides with evaluators blinded to the method. The immunohistochemical intensity and nonspecific background were assessed on pan-cytokeratin and vimentin-stained slides. Overall yield was calculated, and morphologic and immunohistochemical features were compared among paired samples. Technical and cellular features were also described. Agar-based cell blocks and CTBs yielded evaluable sections in 100% (52/52) and 98% (51/52) of the cases, respectively. Cellularity and RBC separation scores were significantly higher in CTBs. Similar staining intensities were observed, and background staining was more frequently seen in pan-cytokeratin-stained ACBs. Only basic materials and equipment were required for both methods. Agar-based cell block preparations were more operator dependent and difficult to standardize, whereas CTBs were easier to prepare, but laboratory processing was more demanding. Both methods can be used to produce good sections for immunohistochemistry staining with no significant differences. Cell tube blocks are beneficial for RBC-rich samples, and little additional training is required to prepare the blocks. Both types of cell blocks are reliable, cost-effective methods that could be introduced in diagnostic laboratories to further characterize canine and feline effusions.

Interobserver Agreement in Pap Stained Smear and Cell Block Preparation in Pleural Fluid Cytology

Interobserver Agreement in Pap Stained Smear and Cell Block Preparation in Pleural Fluid Cytology

Utility of cell block preparation in diagnosis of paediatric abdominal neoplasms.

Paediatric abdominal neoplasms are fairly common. Fine needle aspiration cytology (FNAC) is used for the initial evaluation of paediatric abdominal neoplasms. However, sometimes FNA interpretation can be difficult on limited material, owing to tumour heterogeneity and overlapping features. Therefore, we attempted to obtain additional information using cell block (CB) preparation from residual aspiration material along with immunohistochemistry (IHC) to enhance the diagnostic accuracy. To evaluate the correlation between CB preparation and FNAC in diagnosis of paediatric abdominal tumours with the help of an extended panel of IHC markers and to highlight some of the diagnostic difficulties. A total of 113 cases of paediatric abdominal masses were studied. In addition to routine FNA smears, CBs were prepared from the residual material for IHC analysis as considered appropriate. This study included a total of 113 children with abdominal masses. Histopathology following surgical resection was available in 53 cases. Histology was taken as gold standard to measure the diagnostic accuracy with reference to sensitivity (Sn), specificity (Sp), positive predictive value and negative predictive value. The Sn of FNA alone was 87.5% and Sp was 97.78%. The Sn and Sp increased with use of CB alone and were 100% and 97.78% respectively. The highest Sn and Sp were observed when CB was combined with IHC where both the values were 100%. CB with IHC is a useful adjunct to the routine FNA smears that further contributes to enhance the accuracy of the cytopathological diagnosis and is useful for choosing pre-operative chemotherapeutic regimen.

Utilization of cytology smears improves success rates of RNA-based next-generation sequencing gene fusion assays for clinically relevant predictive biomarkers.

The use of RNA-based next-generation sequencing (NGS) assays to detect gene fusions for targeted therapy has rapidly become an essential component of comprehensive molecular profiling. For cytology specimens, the cell block (CB) is most commonly used for fusion testing; however, insufficient cellularity and/or suboptimal RNA quality are often limiting factors. In the current study, the authors evaluated the factors affecting RNA fusion testing in cytology and the added value of smears in cases with a suboptimal or inadequate CB. A 12-month retrospective review was performed to identify cytology cases that were evaluated by a targeted RNA-based NGS assay. Samples were sequenced by targeted amplicon-based NGS for 51 clinically relevant genes on a proprietary platform. Preanalytic factors and NGS quality parameters were correlated with the results of RNA fusion testing. The overall success rate of RNA fusion testing was 92%. Of the 146 cases successfully sequenced, 14% had a clinically relevant fusion detected. NGS testing success positively correlated with RNA yield (P = .03) but was independent of the tumor fraction, the tumor size, or the number of slides used for extraction. CB preparations were adequate for testing in 45% cases, but the inclusion of direct smears increased the adequacy rate to 92%. There was no significant difference in testing success rates between smears and CB preparations. The success of RNA-based NGS fusion testing depends on the quality and quantity of RNA extracted. The use of direct smears significantly improves the adequacy of cytologic samples for RNA fusion testing for predictive biomarkers.

Validation of PD-L1 Immunohistochemical Testing on Formalin Fixed Paraffin Embedded (FFPE) Cell Block (CB) Preparations

Validation of PD-L1 Immunohistochemical Testing on Formalin Fixed Paraffin Embedded (FFPE) Cell Block (CB) Preparations

Diagnosis of metastatic adenoid cystic carcinoma of the breast on pleural fluid cytology in a 60‐year‐old male

Adenoid cystic carcinoma (AdCC) is a rare triple-negative breast cancer with a favorable prognosis, although cases of distant metastases have been reported. Less than 10 cases of AdCC of the breast in males have been reported. We report a unique presentation of pleural fluid metastasis diagnosed by cytologic evaluation of pleural fluid in a 60-year-old male patient with AdCC of the breast. Careful surveillance and cytologic evaluation of effusion specimens can detect metastases with the aid of cellblock preparations and immunohistochemical stains.

Endoscopic Ultrasound-guided Sampling of Solid Pancreatic Lesions: A Comparative Analysis of 25 Gauge Versus 22 Gauge Core Biopsy Needles.

Pancreatic mass sampling has historically been performed by endoscopic ultrasound-guided fine needle aspiration (EUS-FNA). However, its sensitivity has been reported to be within a wide range, which limits its reliability. Fine needle biopsy (FNB) has been shown to have superior diagnostic performance and is increasingly replacing fine needle aspiration. In FNA, 25 gauge (G) needles appear to outperform 22G. Data comparing these sizes in FNB platforms is limited. We aimed to prospectively compare the performance of 22G and 25G Franseen-tip core biopsy needles in the sampling of solid pancreatic lesions. Patients who underwent EUS-FNB of pancreatic lesions at the Indiana University Hospital using 2 needle sizes: 25G (Study group) and 22G (Control group) using the Acquire needle (Boston Scientific Co., Natick, MA, USA) were enrolled. Needle choice was left to the discretion of the endosonographer. Tissue specimens were evaluated onsite, and underwent touch and smear and cellblock preparation. Specimens were independently evaluated by 2 expert cytopathologists blinded to diagnosis. Cytopathologists assessed cytological yield (on smears) and histological yield (on cellblock) using a validated scoring system reached by a consensus among our cytopathologists as we have previously published. A total of 75 patients (42 males, median=65 years) underwent EUS-FNB during the study period (2017-2018): 50 using 25G and 25 using 22G needle. Diagnostic yield was numerically higher in 25G (98% vs. 88%, p=0.105). Number of passes for smears were similar, however the 25G group required additional passes for cell-block (1.6 vs. 0.4, p=0.001). 25G was used more frequently for pancreatic head and uncinate process sampling (70% vs. 52%, p=0.126). Four patients had self-limited adverse events in the 22G group, but none in the 25G group. We report no difference in the diagnostic yield between 25G FNB vs. 22G sampling device with Franseen style tip, however, the 25G needle use was associated with the need of additional passes to collect a sufficient cell block.

Cell blocks in urine cytopathology: do they add value to the diagnosis? A pilot study

The utility of cell block (CB) preparation is well established in cytopathology. Despite 23.3% of College of American Pathologists-accredited laboratories using CB with liquid-based preparations on urine cytology (UC) cases, there are very few studies on their performance. To determine their usefulness, we conducted a retrospective review of UC cases that received CB. We identified 27 UC cases with ThinPrep (TP) and CB preparation between 2016 and 2020 at our institution. Clinical history and follow-up data were compiled. A blinded review of TP alone, and TP together with CB, was performed by 2 pathologists and 2 cytotechnologists. Diagnoses were rendered in accordance with The Paris System for Reporting Urine Cytology. Blood and acute inflammation were common background elements in cases that received CB preparation. In total, CB upgraded the diagnosis in 7 of 27 cases (26%). The maximum utility of CB preparation was seen in indeterminate cases where 60% (6 of 10) were upgraded, including 71% (5 of 7) of atypical urothelial cells (AUC) and 30% (1 of 3) of suspicious for high-grade urothelial carcinoma (HGUC). One case (1 of 12, 8%) diagnosed as negative for HGUC on TP was diagnosed as low-grade urothelial neoplasia on CB. Our results demonstrate that adjunct use of CB preparation aids in a definitive diagnosis on AUC category and may be helpful in cases with cell clusters or tissue fragments, or cases suspicious for HGUC. Further correlation studies are warranted in this area to expand our knowledge about the utility of CBs in urine cytology.

The diagnostic utility of cell block in fine needle aspiration cytology of palpable breast lesions in a Nigerian tertiary health institution.

Although fine needle aspiration cytology (FNAC) is highly accurate for detecting breast malignancies, concerns remain among cytopathologists about false-positive and false-negative diagnoses. Cell block (CB) preparations have been advocated by some cytopathologists as one of the methods to improve and consolidate the diagnostic accuracy of FNAC. The aim of this study was to determine the diagnostic utility of CB in FNAC of palpable breast lesions among female patients. Following FNA, CBs were prepared using 10% neutral-buffered formalin from the residual breast aspirates of 100 consecutive female patients attending the FNAC clinic. The slides of the conventional smears, CB and excisional biopsies were examined, and results were analysed using the SPSS. Of the 100 patients that had FNAC, 44 (44%) had excisional biopsy performed. An additional 13% diagnostic yield for malignancy was obtained with the use of CB preparations. CB reduced equivocal diagnoses by 25%, corresponding with 90.9% improvement on definitive diagnoses. In our setting, the addition of CB to smear remarkably improved the diagnostic utility of breast FNAC by minimising atypical and suspicious for malignancy diagnostic categories.

The screening value of the numerical control standardized cell block preparation technique in high-grade squamous epithelial lesions of the cervix

The screening value of the numerical control standardized cell block preparation technique in high-grade squamous epithelial lesions of the cervix

Mucinous carcinoma originating in the peritoneum diagnosed by an ascites cell block: a case report

BackgroundPeritoneal carcinoma is a rare disease that is diagnosed and treated in a manner similar to ovarian cancer. In most cases, the histological type is serous carcinoma, and chemotherapy is effective. However, there are a few case reports of mucinous peritoneal carcinoma. We inferred the histological type before surgery using an ascites cell block sample, which was useful for determining the treatment plan.Case presentationOur patient was a 60-year-old Japanese woman. She presented with a feeling of fullness in the abdomen. A computed tomographic scan showed a large quantity of ascitic fluid and thickening of the greater omentum, as well as thickening of the peritoneum at the pouch of Douglas and diaphragm. Hence, peritoneal carcinoma was suspected. The tumor markers carcinoembryonic antigen, cancer antigen 19-9, and cancer antigen 125 were all increased, and no malignant findings were observed in the uterus or ovaries. Cells suggestive of carcinoma were found in the ascitic fluid, and immunostaining by the cell block method suggested the possibility of mucinous carcinoma. The preoperative chemotherapy strategy was changed to short courses, and tumor reduction surgery was planned. Similar to the suspicion before surgery, the pathology results indicated mucinous carcinoma, and the therapeutic effect of chemotherapy was grade 0.ConclusionsDetermining whether peritoneal carcinoma is serous carcinoma is important for therapy and prognostic prediction. In this case, we encountered a patient for whom surgery was chosen because of drug therapy resistance inferred through histological type estimation using the cell block method. Inferring the histological type by cell block preparation is useful for diagnosis and treatment selection.

Enhancing Cell Block Quality- A Comparative Study Of Formalin And Agar-Based Methods

Background: There are not many studies conducted in India to compare cell block preparation methods with reagents and materials that are readily available in all laboratories. This study aimed to standardize and compare two simple cell block techniques, which can be done in low resource settings too. In the study, 35 cases of thyroid, lymph node, and breast were collected for both FNA and cell block preparation for six months.&#x0D; Materials and Methods: There were separate passes given for both methods. A total of seventy cell blocks made using formalin and agar methods of preparation.&#x0D; Results: We compared both the methods on technical and morphological levels. The formalin method was overall easy to perform and was yielding good morphological results in 98% cases, the only drawback being cell loss during handling and processing. While in the agar method, there was almost no cell loss, but it was more technically difficult and yielded poorer morphological results. A scoring system was made for cellularity: no cells = 0, hypo-cellular = 1+, hypo-cellular with tissue fragments = 2+, cellular = 3+.18 A score of 2+ and 3+ was scored by 31/35 formalin blocks and 28/35 agar blocks.&#x0D; Conclusions: The sensitivity of both formalin and agar methods are almost comparable. However, the procedure of the formalin method is far more straightforward and user friendly. Moreover, it also provides a better architectural picture than the agar method.

Implementation of PD-L1 22C3 IHC pharmDxTM in Cell Block Preparations of Lung Cancer: Concordance with Surgical Resections and Technical Validation of CytoLyt® Prefixation

Background: Programmed death ligand-1 (PD-L1) assessed by immunohistochemistry (IHC) is used as biomarker for pembrolizumab therapy in advanced stage lung cancer patients. However, data permitting direct performance comparison between cytology and surgical specimen types are limited since both specimens from a single tumor site are infrequently available. In addition, alcohol fixation used with cytology specimens requires technical validation of the PD-L1 IHC assay before clinical use. We here report our experience with implementation of the PD-L1 22C3 IHC pharmDx^TM assay for cytologic samples at a large tertiary cancer center. Study Design: Archival formalin-fixed (FF), paraffin-embedded cell blocks (CBs) and subsequent lung tumor resections (LTRs) from the same anatomical site were used for a direct comparison of PD-L1 tumor proportion scores (TPSs). TPS values were independently determined by one surgical lung pathologist and two cytopathologists blinded to the specimen pairs. An interim analysis was performed to facilitate the pooling of expertise among observers. After PD-L1 22C3 IHC pharmDx^TM implementation for FF cytology specimens, dual-processed samples were used for a prospective technical validation of CytoLyt® prefixation (CF). Digital image analysis was performed for a subset of dual-processed specimens. Results: Eighty-one CBs and LTRs were included for comparison of the specimen types. PD-L1 assessment in CBs had an accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of 88.9/72.8, 66.7/73.5, 95.2/72.3, 80.0/65.8, and 90.9/79.1% for the ≥50/≥1% cutoff, respectively. The intraclass correlation coefficient was 0.84 (95% confidence interval [CI]: 0.76, 0.90), and it improved after interim analysis (before: 0.79 and after: 0.92). The overall concordance between CF and FF for the categories defined by the ≥50/≥1% cutoff values was 90.4% (95% CI: 79.0, 96.8). Similar assay performance was confirmed by digital analysis. Conclusions: PD-L1 22C3 IHC pharmDx^TM shows good reliability if used with CB preparations. CF does not impact assay results significantly. Clinical validation with outcome data is needed, and digital methods of assessment should be further investigated.

Handling of Thyroid FNA Samples During the COVID-19 Pandemic

Handling of Thyroid FNA Samples During the COVID-19 Pandemic

PD-L1 expression in cell-blocks of non-small cell lung cancer: The impact of prolonged fixation.

In the selection of non-small cell lung cancer (NSCLC) patients for immunotherapy, specimen processed as cell blocks (CBs) may be the only available material to assess PD-L1 expression. Therefore, optimal CB preparation becomes paramount. In this context, here we assessed whether inadequate fixation time might be one of the pre-analytical factors affecting PD-L1 expression. Ex vivo CBs from placental (n = 3) and NSCLC (n = 8) resection specimens were obtained. PD-L1 staining was performed on CBs prepared at increasing fixation times (12 hours, 48 hours, 72 hours, 96 hours, 168 hours and 504 hours) using the companion diagnostic SP263 Assay and a validated 22C3 laboratory developed test (LDT). Staining intensity and percentage of positive cells were evaluated. All placental CBs showed moderate to strong PD-L1 positivity in most cells, regardless of the fixation time. Likewise, the percentage of SP263-stained NSCLC cells was similar at all fixation times except for one case, which showed less intense SP263 staining at 168 hours. Conversely, in 5/8 cases, the 22C3 LDT percentage of positive cells and staining intensity decreased at 168 hours and 504 hours. Our results show that fixation time influences the performance of 22C3 LDT on CBs. Thus, we recommend that the fixation time of cytological materials be carefully checked, especially when PD-L1 testing is delayed until the oncology request. Indeed, delays in tissue processing and paraffin embedding may lead to sub-optimal performance of PD-L1 staining on CBs.

Cytopathological correlation of cell block in ultrasound guided fine needle aspiration cytology

Introduction: Despite many advantages, at times FNA does not yield enough information for precise diagnosis, the material remaining in the hub or extra material can be used in Cell Block preparation. Present study was done to assess utility of USG guided FNAC with CB preparation in increasing sensitivity and diagnostic accuracy of deep-seated lesions. Methodology: One-year cross sectional study carried out in a tertiary hospital, on 50 patients referred for USG guided FNAC of deep-seated lesions after considering inclusion/exclusion criteria. Consent and detailed clinical history were taken. After making conventional FNAC smears, Fixed sedimentation method was used for cell block preparation by flushing residual material in needle hub and syringe in 10% buffered formalin. Centrifugation at 3000 rpm was done for 10 minutes to form pellets, supernatant was decanted, deposit was wrapped in filter paper and processed as regular surgical tissue. FNAC smears and Cell block sections were examined and results were correlated with histopathology.Results: Of the 50 cases, Sensitivity, specificity, and diagnostic accuracy of deep-seated lesions considering histopathology as gold standard was 86.96%, 100%, 88.0% for FNAC and 95.65%, 100%, 96.0% for Cell block and 97.83%, 100%, 98.0% for FNAC+ CB combined respectively.Conclusion: Cell blocks as an adjuvant to FNAC smears is highly effective technique in cytology for definitive diagnosis.