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Purpose: The purpose of this study was to examine the diagnostic accuracy of the cell block (CB) method and clinical features affecting it in patients with vitreoretinal lymphoma (VRL). Methods: This study enrolled 38 eyes in 33 VRL patients, and 7 eyes in 7 patients with idiopathic uveitis who underwent diagnostic vitrectomy. Medical records including the results of CB cytology, interleukin (IL)-10/-6 concentrations, and immunoglobulin heavy chain gene (IgH) rearrangement were retrospectively searched. Results: Patients with VRL comprised 16 women and 17 men, and the age of onset ranged from 44 to 85 years (mean: 70 years). CB preparations detected large malignant cells in 35 eyes (92%), whereas the other 3 VRL eyes were negative. Two of the latter three eyes showed subretinal infiltrates, which existed in 7 of 35 CB-positive eyes. Intravitreal IL-10 and -6 concentrations were 1866 ± 4088 pg/mL and 98 ± 139 pg/mL, respectively, and the rate of IL-10/-6 >1 was 86.9%. The presence of IgH monoclonality was 63.2%. In patients with uveitis, CB specimens revealed no atypical but small inflammatory cells. IL-6 concentration was 311.1 ± 240 pg/mL, whereas IL-10 was undetectable in six eyes, and the IL-negative rate was 85.7%. Six eyes (85.7%) with uveitis showed no IgH monoclonality. Conclusions: Diagnostic accuracy of CB preparations in VRL could achieve an equivalent outcome to IL ratio calculation and IgH monoclonality detection. The appearance of subretinal infiltrates may diminish the CB positivity.

Background: Distinction between reactive mesothelial cells, malignant mesothelioma and carcinoma is challenging in both biopsy and cytologic material. This study was conducted to differentiate benign/ reactive mesothelial proliferation from malignant mesothelial proliferations and metastatic adenocarcinoma by using immunohistochemical (IHC) markers Desmin (DES), Epithelial membrane antigen (EMA) and Calretinin (CAL) in pleural uid cell block (CB) preparations. A two year descriptive study (Oct.2016- Sept.2018). 46 pleural uids samples sentMethods : to the Dept. of Pathology, RIMS for routine examination and histopathological examination by CB preparation were studied using IHC markers EMA,DES and CAL following H & E stain. Out of 46 cases, 9(19.6%) cases were diagnosed as benign, 23(50.0%) as reactive andResults: 14(30.4%) as adenocarcinoma on H & E section by CB preparations within an age range 34 to 80 years (Mean±SE, 60.32±12.13). Following IHC staining with EMA, DES & CAL, 11(23.9%) cases were conrmed as benign, 17(37.0%) as reactive, 16(34.8%) as adenocarcinoma and 2(4.3%) cases as malignant mesothelioma. This study showed that EMA, DES and CAL helpful in conrming benign or reactive mesothelialConclusions: and malignant mesothelial with epithelial cells which will be helpful in providing appropriate diagnosis in difcult cases and provide better patient management.

BACKGROUND Inconclusive cytomorphology often results due to failure to distinguish between adenocarcinoma cells from benign, reactive, atypical mesothelial cells in effusion specimens. To resolve such dilemmas, auxiliary techniques like immunohistochemistry were utilised to reach a definitive diagnosis for better treatment and management of patients. We wanted to compare cytodiagnosis achieved on cell block preparations with the cytodiagnosis on conventional smear and perform immunohistochemistry for epithelial membrane antigen (EMA), calretinin, desmin, vimentin and E-cadherin on cell block preparation of the fluids in cases of indistinguishable cytomorphology of adenocarcinoma and reactive, atypical, and benign mesothelial hyperplasia. METHODS The immunohistochemical markers namely EMA, calretinin, vimentin, desmin and Ecadherin were applied on cell blocks employing streptavidin-biotin method. Immunohistochemistry was interpreted by giving scores to the percentage of stained cells. RESULTS EMA and E-cadherin had 100 % sensitivity in diagnosing adenocarcinoma whereas calretinin, vimentin and desmin were 100 % sensitive in diagnosing reactive, atypical mesothelial carcinoma on the cell block preparations. CONCLUSIONS Immunocytochemistry of fluid should be carried out on the cell block preparation where cytological diagnosis on conventional smear and cell block fails to detect malignant cells in the preparation. KEY WORDS Cell Block, Adenocarcinoma, Mesothelial Cells, Immunohistochemistry, EMA, Calretinin, Vimentin, Desmin, E-Cadherin

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 pharmDxTM 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 pharmDxTM 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 pharmDxTM 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.

Objective Primary retroperitoneal involvement of lymphoproliferative disorders is rare. Diffuse large B cell lymphoma (DLBCL) in the retroperitoneum has been scarcely reported and the diagnostic workup is challenging due to difficulties in access for tissue acquisition. Materials and methods A 73-year-old female presented with epigastric pain and nausea. A computed tomography scan revealed retroperitoneal lymphadenopathy. An EUS-FNA was performed and a cytological diagnosis of lymphomatous disorder was made on direct smears .A panel of immunocytochemical markers was applied on cell block preparations and revealed a EBV related DLBCL (CD20 positive, PAX-5 positive, CD3 negative, MUM-1 positive, BCL-2 positive, BCL-6 positive, CD10 positive, MIB-1 positive, and LMP-1 positive). Conclusion Our case illustrates the utility of EUS-FNA in the work-up of retroperitoneal lymphadenopathy and the importance of cell block processing in the diagnostic approach of EBV related DLBCL.

Background The identification of BRAF mutations in thyroid cancer has prognostic and therapeutic implications. Although the gold standard for identifying BRAF mutations is molecular testing, the ability to perform BRAF p.V600E immunostaining on fine-needle aspiration (FNA) samples can facilitate the rapid triaging of patients to treatment options. Methods A total of 50 thyroid carcinoma FNA samples, including papillary (29 samples), poorly differentiated (10 samples), anaplastic (9 samples), and Hurthle cell (2 samples) carcinomas, with a known BRAF p.V600E mutation status were selected for the current study. Immunostaining was performed on smears and cell block sections using an anti-BRAF p.V600E antibody (clone VE1). The results were compared with the known mutation status obtained by molecular testing and/or immunostaining of surgical pathology material from the same patient. Results Of the total of 50 cases, 26 cases had smears available for the evaluation of BRAF p.V600E immunostaining; positive immunostaining was noted in 16 samples and negative immunostaining was noted in 4 samples, whereas 6 cases were equivocal. Of the 34 cases for which cell blocks were available for evaluation, BRAF p.V600E immunostaining was positive in 17 cases, negative in 16 cases, and equivocal in 1 case. The overall sensitivity and specificity of BRAF p.V600E immunostaining on the cell block preparation was 94.4% and 100%, respectively, whereas for the smears it was 80% and 63.6%, respectively. Conclusions BRAF p.V600E immunostaining can be performed reliably on thyroid FNA cell block preparations. However, false-positive results on direct smears limit their utility and therefore need to be interpreted with caution. Cancer Cytopathol 2018;126:406-13. © 2018 American Cancer Society.

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 pharmDxTM 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 pharmDxTM 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 pharmDxTM 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.

Studies using the cell-block technique for bone marrow (BM) analysis are limited in the veterinary literature. This work assessed whether the histopathological analysis of canine BM was feasible using cell-block cytoinclusions prepared from fine-needle sternal aspirate samples. Eight clinically healthy young-to-middle-aged dogs underwent both fine-needle sternal aspiration for BM cell-block (BM-Cb) processing and iliac-crest BM core biopsy (BM-B). Histopathologic parameters were compared between the 2 methods. There were no statistically significant histopathological differences between hematopoietic tissue areas (P = .6294) in the BM-Cb and BM-B sections, and they had similar microscopic characteristics and microarchitecture. Cellularity and reticulin-fiber staining were equivalent in the BM-Cb and BM-B preparations in 87.5% (7/8) and 100% (8/8) of the sections, respectively. However, the quantitative results of the megakaryocytic series differed between BM-Cb and BM-B in 37.5% (3/8) of the sections, and the myeloid:erythroid (M:E) ratios differed between the 2 methods in 25% (2/8). These preliminary data indicate that cell-block preparations made from sternal fine-needle aspiration samples warrant continued evaluation in a larger number of animals, including those with various diseases affecting the bone marrow.

Introduction The diagnosis of pancreatic ductal adenocarcinoma (PDA) on endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) material is often challenging. An immunohistochemical (IHC) panel may help establish the diagnosis of PDA in cases limited by sample size or ambiguous cytology. S100P, IMP3, and SMAD4 are 3 IHC markers that have shown promise as individual markers for PDA that have never been tested together as a panel. In this study, we evaluated the individual and combined efficacy of S100P, IMP3, and SMAD4 for the detection of PDA. Materials and Methods S100P, IMP3, and SMAD4 IHC staining was performed on cell blocks (CBs) procured from pancreatic EUS-FNA procedures. The cohort included CBs that were diagnostic for PDA (n = 35), suspicious but nondiagnostic for PDA (n = 2), as well as CBs with benign pancreatic ductal epithelium (n = 12) and benign reactive pancreatic ductal epithelium (n = 18). A positive result for IMP3 and S100P was defined as moderate or strong staining of >10% of ductal cells. Complete lack of SMAD4 nuclear staining was considered a positive result—any nuclear SMAD4 staining was considered a negative result. Results Two and 3 IHC marker panels were almost always more specific than individual IHC markers. Positivity for at least 2 of 3 IHC markers was a sensitive (91.89%) and highly specific (100%) marker of PDA. Conclusions The 3 IHC marker panel composed of S100P, IMP3, and SMAD4 is highly specific for PDA. Future studies should evaluate efficacy in a cohort with more atypical and suspicious cases.

Background: Cell Block (CB) procedures have now become an established part of cytological diagnostics because of its pivotal role in diagnosis and ancillary studies. Hence the present study was undertaken to emphasize the role of CB technique over Conventional Smear (CS) in serous effusions and to compare the Plasma-Thrombin (PT) block to Formalin Method block (FM) in assessment of morphological preservation and cellularity. Aim: To obtain simple , cost effective and ideal CB preparation where in maximal number of cells are displayed within a small area Methods: The sample was divided into three Parts(A,B,C). After centrifugation of all three parts of sample at 3000rpm for 15min – Part A sediment was used to prepare two CS for Papanicolaou (PAP) and May Grunwald Giemsa(MGG) stains. Part B sediment was subjected for 1hr and 24hr fixation in 1:1 solution of 5ml ethylalcohol and 10% formalin. To the Part C sediment 2drops of finger prick plain blood was added, mixed well and allowed to clot. The sediment of Part B and the clot of Part C were then processed for paraffin embedding. Result: 110 fresh effusion samples were evaluated for cellularity retention of architectural patterns and volume of background. FM block’s were inconclusive in 12 cases due to low cellularity. PT block’s were all evaluable with best preservation of architecture and pale background. Conclusion: The CB technique revealed better architectural patterns and increased the sensitivity of cytodiagnosis. PT block’s had sufficient to abundant cellularity with evenly distributed cells in small area. PT preparation is simple and cost effective. DOI: 10.21276/APALM.1432

Objective Atypical squamous cells of undetermined significance (ASCUS) and atypical glandular cells (AGC) reflect cellular abnormalities insufficient for clear diagnosis. We used cell pellets obtained from liquid-based cytology (LBC) to prepare cell blocks (CB) and clarify the initial diagnosis of ASCUS and AGC. Study Design A total of 393 CBs with initial diagnosis of ASCUS or AGC were processed. Of those, 305 of the ASCUS and 20 of the AGC had adequate specimens. We compared results of CBs prepared from ASCUS and AGC to determine which had higher frequencies of higher-grade lesions. Results A majority of specimens (83%) were adequate for evaluation. Compared with the initial diagnosis, 14% (42/305) of ASCUS were diagnosed with low-grade squamous intraepithelial lesion (LSIL) in CB, while 10% (2/20) of AGC were diagnosed with LSIL or adenocarcinoma. No statistical relationship between the initial diagnosis of ASCUS and AGC and results of higher-grade lesions in CB is evident as determined by p value greater than 0.05 (p = 0.228). Conclusions CBs prepared from Liqui-PREP cell pellets are, in most cases, assessable and can be useful as an adjunctive test to help clarify the initial diagnosis of ASCUS and AGC. Diagn. Cytopathol. 2017;45:520–525. © 2017 Wiley Periodicals, Inc.

Myofibroblastoma (MFB) is a benign tumor of the mammary stroma with predominant myofibroblastic differention. The cytologic reports of MFB are very few in the available literature. From the cytodiagnostic point of view about 21 cases of MFB with cytological evaluation by fine-needle aspiration cytology (FNAC) have been encountered in the English-language literature: A 35-year-old woman presented with lump in the left breast. FNAC showed mild degree of pleomorphism with occasional groups with fibrous stroma and tumoral cells. A few benign epithelial cell groups were seen. Hematoxylin-eosin-stained sections of cell block preparation from the aspirate showed fascicles of spindle cells forming whorl structures. Three months later, excision biopsy was performed. The diagnosis was a classic variant of MFB. On immunohistochemical examination, sections were found to be highly positive for vimentin, CD34, and bcl- 2. In light of these findings, cell block material was retrospectively reviewed both morphologically and immunhistochemically. The findings of resection and cell block material were found to be very similar. MFB may cause a potential diagnostic pitfall while interpreting FNAC due to its wide differential diagnosis spectrum. We concluded that cytology and cell block findings complement each other. Diagn. Cytopathol. 2016;44:1064-1069. © 2016 Wiley Periodicals, Inc.

Objectives Programmed death-ligand 1 (PD-L1) is an emerging molecular target in anticancer therapy, most notably non–small cell lung cancers. PD-L1 expression in pancreatic adenocarcinomas (PDAs) on surgical specimens is highly variable, with 10% to 60% of tumors showing expression. PD-L1 expression in PDA on endoscopic ultrasound-guided fine needle aspirations (EUS-FNAs) has been rarely studied. Methods Formalin-fixed, paraffin embedded (FFPE) cell blocks (CBs) from 65 EUS-FNA samples of 55 patients, with a diagnosis of PDA, with at least 20% tumor cellularity were retrieved. The cell blocks were originally fixed in CytoRich fixative. Immunohistochemical staining (IHC) for PD-L1 was performed using the M365329-1 (22C3) clone according to the manufacturer’s approved protocol and optimized for the fixation method using appropriate controls. A combined positive score (CPS) defined per CAP guidelines as the total number of positive tumor cells and inflammatory cells as a percentage of the total number of tumor cells was assessed for each case and was grouped as 20. Results Twenty-five samples (38%) from 21 patients showed immunoreactivity to PD-L1, with 21 (32%) having a CPS of Conclusion Immunohistochemical analysis for PD-L1 is feasible on EUS-FNA CB samples when optimized and validated for the fixation method using appropriate controls. PD-L1 expression is seen in only a minority of PDAs, albeit with a very low CPS. The potential role of PD-L1 as a prognostic marker for disease progression needs further exploration.

Objectives Immunohistochemistry especially for biomarkers such as PD-L1 in personalized medicine is increasingly being performed on cell blocks from cytopathology samples. We evaluated the effects of staining cell blocks (CBs) from different cytologic fixatives using four different commercially available PD-L1 antibody clones. Methods PD-L1 immunohistochemistry (IHC) using four different commercially available clones was performed on eight cell blocks processed from two different fixatives and also on eight formalin-fixed tissues. The clones used were 22C3 at 1:50 dilution and 1:100 dilution, SP263, SP142, and E1L3N. The cell block (CB) samples contained cells that strongly express PD-L1 and either fixed in CytoLyt (methanol-based fixative, n = 4) or CytoRich (ethanol-formalin-based fixative, n = 4). Formalin-fixed control tissues (tonsillar tissue, n = 4; stomach, n = 4) were also included. Results Expression for PD-L1 was noted on all the CytoRich fixed cell blocks (n = 4) for all four antibody clones, with the intensity and distribution of expression comparable to the formalin fixed tissues (n = 8). However, consistent absence of expression for PD-L1 was noted on all the CytoLyt fixed cell blocks (n = 4) for all four antibody clones. Conclusion The results of this pilot study demonstrate that PD-L1 IHC on cell blocks is feasible. However, there is a need to validate IHC protocols according to specific fixation methods.