The Experts below are selected from a list of 4848 Experts worldwide ranked by ideXlab platform
Russell Broaddus - One of the best experts on this subject based on the ideXlab platform.
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salvaging the Supernatant next generation cytopathology for solid tumor mutation profiling
Modern Pathology, 2018Co-Authors: Sinchita Roychowdhuri, Meenakshi Mehrotra, Ana Maria Bolivar, Brette Hannigan, Stephanie Zalles, Wenrui Ye, Dzifa Y Duose, Rashmi Kanagalshamanna, Bedia A Barkoh, Russell BroaddusAbstract:With the expanding role of targeted therapy in patients with solid tumors, pathologists face the daunting task of having to maximize limited volume tissue obtained by fine needle aspiration for a variety of molecular tests. While most molecular studies on fine needle aspiration samples have been reported using cellular material, recent studies have shown that a substantial amount of DNA can be retrieved from the Supernatant Fluid of aspirate needle rinses after cell pelleting for cytospin or cell block preparations. In routine clinical workflow, the Supernatant is discarded; however this Fluid may provide a complementary source of DNA for tumor mutational profiling. In this study, we evaluated the post-centrifuged Supernatant from 25 malignant and 10 benign fine needle aspiration needle rinses. The mean and median DNA yields from the Supernatants were 445 ng and 176.4 ng (range, 15.1–2958 ng), respectively. Next generation sequencing using the Ion AmpliSeq Cancer Hotspot Panel v2 detected somatic mutations in all 25 malignant samples. No mutations were detected in any of the benign samples tested. When available, mutations detected in the Supernatant Fluid were compared to the next generation sequencing analysis performed on a prior or concurrent surgical specimen from the same patient and showed 100% concordance. In a subset of cases (n = 19) mutations in EGFR, KRAS, BRAF, PIK3CA, and NRAS were successfully confirmed by droplet digital PCR, providing an orthogonal platform for mutation analysis. In summary, in this study we show that post centrifuged Supernatants from fine needle aspiration needle rinses can provide a robust substrate for expanded mutation profiling by next generation sequencing, as well as hotspot mutation testing by droplet digital PCR. The ability to detect somatic mutations from otherwise discarded Supernatant Fluids offers the ability to triage and effectively utilize limited volume fine needle aspiration samples when multiple molecular tests are requested, without the need to re-biopsy for additional tissue samples.
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salvaging the Supernatant next generation cytopathology for solid tumor mutation profiling
Modern Pathology, 2018Co-Authors: Sinchita Roychowdhuri, Meenakshi Mehrotra, Ana Maria Bolivar, Brette Hannigan, Stephanie Zalles, Wenrui Ye, Dzifa Y Duose, Rashmi Kanagalshamanna, Bedia A Barkoh, Russell BroaddusAbstract:With the expanding role of targeted therapy in patients with solid tumors, pathologists face the daunting task of having to maximize limited volume tissue obtained by fine needle aspiration for a variety of molecular tests. While most molecular studies on fine needle aspiration samples have been reported using cellular material, recent studies have shown that a substantial amount of DNA can be retrieved from the Supernatant Fluid of aspirate needle rinses after cell pelleting for cytospin or cell block preparations. In routine clinical workflow, the Supernatant is discarded; however this Fluid may provide a complementary source of DNA for tumor mutational profiling. In this study, we evaluated the post-centrifuged Supernatant from 25 malignant and 10 benign fine needle aspiration needle rinses. The mean and median DNA yields from the Supernatants were 445 ng and 176.4 ng (range, 15.1–2958 ng), respectively. Next generation sequencing using the Ion AmpliSeq Cancer Hotspot Panel v2 detected somatic mutations in all 25 malignant samples. No mutations were detected in any of the benign samples tested. When available, mutations detected in the Supernatant Fluid were compared to the next generation sequencing analysis performed on a prior or concurrent surgical specimen from the same patient and showed 100% concordance. In a subset of cases (n = 19) mutations in EGFR, KRAS, BRAF, PIK3CA, and NRAS were successfully confirmed by droplet digital PCR, providing an orthogonal platform for mutation analysis. In summary, in this study we show that post centrifuged Supernatants from fine needle aspiration needle rinses can provide a robust substrate for expanded mutation profiling by next generation sequencing, as well as hotspot mutation testing by droplet digital PCR. The ability to detect somatic mutations from otherwise discarded Supernatant Fluids offers the ability to triage and effectively utilize limited volume fine needle aspiration samples when multiple molecular tests are requested, without the need to re-biopsy for additional tissue samples.
Sydney D Finkelstein - One of the best experts on this subject based on the ideXlab platform.
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the value of mutational profiling of the cytocentrifugation Supernatant Fluid from fine needle aspiration of pancreatic solid mass lesions
Modern Pathology, 2014Co-Authors: Georgios Deftereos, Sydney D Finkelstein, Sara A Jackson, E M Ellsworth, Uma Krishnamurti, Yulin Liu, Jan F Silverman, Candy R Binkert, Beth A Ujevich, Alok MohantyAbstract:Fine-needle aspiration (FNA) of pancreatic solid masses can be significantly impacted by sampling variation. Molecular analysis of tumor DNA can be an aid for more definitive diagnosis. The aim of this study was to evaluate how molecular analysis of the cell-free cytocentrifugation Supernatant DNA can help reduce sampling variability and increase diagnostic yield. Twenty-three FNA smears from pancreatic solid masses were performed. Remaining aspirates were rinsed for preparation of cytocentrifuged slides or cell blocks. DNA was extracted from Supernatant Fluid and assessed for DNA quantity spectrophotometrically and for amplifiability by quantitative PCR (qPCR). Supernatants with adequate DNA were analyzed for mutations using PCR/capillary electrophoresis for a broad panel of markers (KRAS point mutation by sequencing, microsatellite fragment analysis for loss of heterozygosity (LOH) of 16 markers at 1p, 3p, 5q, 9p, 10q, 17p, 17q, 21q, and 22q). In selected cases, microdissection of stained cytology smears and/or cytocentrifugation cellular slides were analyzed and compared. In all, 5/23 samples cytologically confirmed as adenocarcinoma showed detectable mutations both in the microdissected slide-based cytology cells and in the cytocentrifugation Supernatant. While most mutations detected were present in both microdissected slides and Supernatant Fluid specimens, the latter showed additional mutations supporting greater sensitivity for detecting relevant DNA damage. Clonality for individual marker mutations was higher in the Supernatant Fluid than in microdissected cells. Cytocentrifugation Supernatant Fluid contains levels of amplifiable DNA suitable for mutation detection and characterization. The finding of additional detectable mutations at higher clonality indicates that Supernatant Fluid may be enriched with tumor DNA. Molecular analysis of the Supernatant Fluid could serve as an adjunct method to reduce sampling variability and increase diagnostic yield, especially in cases with a high clinical suspicion for malignancy and limited number of atypical cells in the smears.
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molecular analysis of pancreaticobiliary fine needle aspirate duct brushing cytocentrifugation Supernatant Fluid
Journal of Clinical Oncology, 2012Co-Authors: Sydney D Finkelstein, E M Ellsworth, Max C Schmidt, Valerie Stearns, Dennis M SmithAbstract:e21121 Background: Reliance on minimally invasive sampling techniques (fine needle aspiration (FNA), duct brushing) continues to increase for pancreaticobiliary (PB) pre-cancer/cancer diagnosis. Fu...
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molecular analysis of centrifugation Supernatant Fluid from pancreaticobiliary duct samples can improve cancer detection
Acta Cytologica, 2012Co-Authors: Sydney D Finkelstein, Marluce Bibbo, David E Loren, Ali A Siddiqui, Charalambos C Solomides, Thomas E Kowalski, Eric EllsworthAbstract:Objective: We aimed to supplement microscopic examination of biliary cytobrush specimens to improve sensitivity by mutational profiling of: (1) selected cells microdissected from cytology slides and (2) corresponding cell-free DNA in residual Supernatant Fluid. Study Design: From 43 patients with brushings of bile or pancreatic duct strictures, DNA was extracted from microdissected cells and 1–2 ml of cytocentrifugation Supernatant Fluid. Mutational analysis targeted 17 genomic sites associated with pancreaticobiliary cancer, including sequencing for KRAS point mutation and loss of heterozygosity (LOH) analysis of microsatellites located at 1p, 3p, 5q, 9p, 10q, 17p, 17q, 21q, and 22q. Results: Mutations were found in 25/28 patients with malignancy, and no mutations were found in 5/5 patients with benign surgical results. The cell-free Supernatant Fluid generally contained higher levels and quality of DNA, resulting in increased detection of mutations in most patients. KRAS mutations only occurred in patients with pancreatic cancer. Mutational profiling of Supernatant Fluid specimens resulted in high sensitivity and specificity for malignancy, improving the detection of malignancy over cytology alone. Conclusion: Brush cytology specimens yielded Supernatant Fluid enriched with DNA, probably from actively proliferating cells. Mutational profiling can enhance the cytologic evaluation and characterization of specimens suspected to contain pancreatic or bile duct cancer.
Sinchita Roychowdhuri - One of the best experts on this subject based on the ideXlab platform.
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salvaging the Supernatant next generation cytopathology for solid tumor mutation profiling
Modern Pathology, 2018Co-Authors: Sinchita Roychowdhuri, Meenakshi Mehrotra, Ana Maria Bolivar, Brette Hannigan, Stephanie Zalles, Wenrui Ye, Dzifa Y Duose, Rashmi Kanagalshamanna, Bedia A Barkoh, Russell BroaddusAbstract:With the expanding role of targeted therapy in patients with solid tumors, pathologists face the daunting task of having to maximize limited volume tissue obtained by fine needle aspiration for a variety of molecular tests. While most molecular studies on fine needle aspiration samples have been reported using cellular material, recent studies have shown that a substantial amount of DNA can be retrieved from the Supernatant Fluid of aspirate needle rinses after cell pelleting for cytospin or cell block preparations. In routine clinical workflow, the Supernatant is discarded; however this Fluid may provide a complementary source of DNA for tumor mutational profiling. In this study, we evaluated the post-centrifuged Supernatant from 25 malignant and 10 benign fine needle aspiration needle rinses. The mean and median DNA yields from the Supernatants were 445 ng and 176.4 ng (range, 15.1–2958 ng), respectively. Next generation sequencing using the Ion AmpliSeq Cancer Hotspot Panel v2 detected somatic mutations in all 25 malignant samples. No mutations were detected in any of the benign samples tested. When available, mutations detected in the Supernatant Fluid were compared to the next generation sequencing analysis performed on a prior or concurrent surgical specimen from the same patient and showed 100% concordance. In a subset of cases (n = 19) mutations in EGFR, KRAS, BRAF, PIK3CA, and NRAS were successfully confirmed by droplet digital PCR, providing an orthogonal platform for mutation analysis. In summary, in this study we show that post centrifuged Supernatants from fine needle aspiration needle rinses can provide a robust substrate for expanded mutation profiling by next generation sequencing, as well as hotspot mutation testing by droplet digital PCR. The ability to detect somatic mutations from otherwise discarded Supernatant Fluids offers the ability to triage and effectively utilize limited volume fine needle aspiration samples when multiple molecular tests are requested, without the need to re-biopsy for additional tissue samples.
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salvaging the Supernatant next generation cytopathology for solid tumor mutation profiling
Modern Pathology, 2018Co-Authors: Sinchita Roychowdhuri, Meenakshi Mehrotra, Ana Maria Bolivar, Brette Hannigan, Stephanie Zalles, Wenrui Ye, Dzifa Y Duose, Rashmi Kanagalshamanna, Bedia A Barkoh, Russell BroaddusAbstract:With the expanding role of targeted therapy in patients with solid tumors, pathologists face the daunting task of having to maximize limited volume tissue obtained by fine needle aspiration for a variety of molecular tests. While most molecular studies on fine needle aspiration samples have been reported using cellular material, recent studies have shown that a substantial amount of DNA can be retrieved from the Supernatant Fluid of aspirate needle rinses after cell pelleting for cytospin or cell block preparations. In routine clinical workflow, the Supernatant is discarded; however this Fluid may provide a complementary source of DNA for tumor mutational profiling. In this study, we evaluated the post-centrifuged Supernatant from 25 malignant and 10 benign fine needle aspiration needle rinses. The mean and median DNA yields from the Supernatants were 445 ng and 176.4 ng (range, 15.1–2958 ng), respectively. Next generation sequencing using the Ion AmpliSeq Cancer Hotspot Panel v2 detected somatic mutations in all 25 malignant samples. No mutations were detected in any of the benign samples tested. When available, mutations detected in the Supernatant Fluid were compared to the next generation sequencing analysis performed on a prior or concurrent surgical specimen from the same patient and showed 100% concordance. In a subset of cases (n = 19) mutations in EGFR, KRAS, BRAF, PIK3CA, and NRAS were successfully confirmed by droplet digital PCR, providing an orthogonal platform for mutation analysis. In summary, in this study we show that post centrifuged Supernatants from fine needle aspiration needle rinses can provide a robust substrate for expanded mutation profiling by next generation sequencing, as well as hotspot mutation testing by droplet digital PCR. The ability to detect somatic mutations from otherwise discarded Supernatant Fluids offers the ability to triage and effectively utilize limited volume fine needle aspiration samples when multiple molecular tests are requested, without the need to re-biopsy for additional tissue samples.
Alok Mohanty - One of the best experts on this subject based on the ideXlab platform.
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the value of mutational profiling of the cytocentrifugation Supernatant Fluid from fine needle aspiration of pancreatic solid mass lesions
Modern Pathology, 2014Co-Authors: Georgios Deftereos, Sydney D Finkelstein, Sara A Jackson, E M Ellsworth, Uma Krishnamurti, Yulin Liu, Jan F Silverman, Candy R Binkert, Beth A Ujevich, Alok MohantyAbstract:Fine-needle aspiration (FNA) of pancreatic solid masses can be significantly impacted by sampling variation. Molecular analysis of tumor DNA can be an aid for more definitive diagnosis. The aim of this study was to evaluate how molecular analysis of the cell-free cytocentrifugation Supernatant DNA can help reduce sampling variability and increase diagnostic yield. Twenty-three FNA smears from pancreatic solid masses were performed. Remaining aspirates were rinsed for preparation of cytocentrifuged slides or cell blocks. DNA was extracted from Supernatant Fluid and assessed for DNA quantity spectrophotometrically and for amplifiability by quantitative PCR (qPCR). Supernatants with adequate DNA were analyzed for mutations using PCR/capillary electrophoresis for a broad panel of markers (KRAS point mutation by sequencing, microsatellite fragment analysis for loss of heterozygosity (LOH) of 16 markers at 1p, 3p, 5q, 9p, 10q, 17p, 17q, 21q, and 22q). In selected cases, microdissection of stained cytology smears and/or cytocentrifugation cellular slides were analyzed and compared. In all, 5/23 samples cytologically confirmed as adenocarcinoma showed detectable mutations both in the microdissected slide-based cytology cells and in the cytocentrifugation Supernatant. While most mutations detected were present in both microdissected slides and Supernatant Fluid specimens, the latter showed additional mutations supporting greater sensitivity for detecting relevant DNA damage. Clonality for individual marker mutations was higher in the Supernatant Fluid than in microdissected cells. Cytocentrifugation Supernatant Fluid contains levels of amplifiable DNA suitable for mutation detection and characterization. The finding of additional detectable mutations at higher clonality indicates that Supernatant Fluid may be enriched with tumor DNA. Molecular analysis of the Supernatant Fluid could serve as an adjunct method to reduce sampling variability and increase diagnostic yield, especially in cases with a high clinical suspicion for malignancy and limited number of atypical cells in the smears.
Eric Ellsworth - One of the best experts on this subject based on the ideXlab platform.
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mutational analysis of cytocentrifugation Supernatant Fluid from pancreatic solid mass lesions
Diagnostic Cytopathology, 2014Co-Authors: D Sydney M D Finkelstein, Marluce Bibbo, Ali A Siddiqui, E Thomas M D Kowalski, E David M D Loren, M Charalambos D Solomides, Eric EllsworthAbstract:Diagnosis of fine-needle aspirations of pancreatic solid masses is complicated by many factors that keep its false-negative rate high. Our novel approach analyzes cell-free cytocentrifugation Supernatant, currently a discarded portion of the specimen. Supernatant and cytology slides were collected from 25 patients: 11 cases with confirmed outcome [five positive (adenocarcinoma) and six negative (inflammatory states)], plus 14 without confirmed outcomes. Slides were microdissected, DNA was extracted from microdissections and corresponding Supernatants, and all were analyzed for KRAS point mutation and loss of heterozygosity. Notably, higher levels of free DNA were found in Supernatants than in corresponding microdissected cells. Supernatants contained sufficient DNA for mutational profiling even when samples contained few to no cells. Mutations were present in 5/5 malignancies and no mutations were present in inflammatory states. In conclusion, these findings support using Supernatant for mutational genotyping when diagnostic confirmation is required for pancreatic solid masses. Diagn. Cytopathol. 2014;42:719–725. © 2013 Wiley Periodicals, Inc.
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molecular analysis of centrifugation Supernatant Fluid from pancreaticobiliary duct samples can improve cancer detection
Acta Cytologica, 2012Co-Authors: Sydney D Finkelstein, Marluce Bibbo, David E Loren, Ali A Siddiqui, Charalambos C Solomides, Thomas E Kowalski, Eric EllsworthAbstract:Objective: We aimed to supplement microscopic examination of biliary cytobrush specimens to improve sensitivity by mutational profiling of: (1) selected cells microdissected from cytology slides and (2) corresponding cell-free DNA in residual Supernatant Fluid. Study Design: From 43 patients with brushings of bile or pancreatic duct strictures, DNA was extracted from microdissected cells and 1–2 ml of cytocentrifugation Supernatant Fluid. Mutational analysis targeted 17 genomic sites associated with pancreaticobiliary cancer, including sequencing for KRAS point mutation and loss of heterozygosity (LOH) analysis of microsatellites located at 1p, 3p, 5q, 9p, 10q, 17p, 17q, 21q, and 22q. Results: Mutations were found in 25/28 patients with malignancy, and no mutations were found in 5/5 patients with benign surgical results. The cell-free Supernatant Fluid generally contained higher levels and quality of DNA, resulting in increased detection of mutations in most patients. KRAS mutations only occurred in patients with pancreatic cancer. Mutational profiling of Supernatant Fluid specimens resulted in high sensitivity and specificity for malignancy, improving the detection of malignancy over cytology alone. Conclusion: Brush cytology specimens yielded Supernatant Fluid enriched with DNA, probably from actively proliferating cells. Mutational profiling can enhance the cytologic evaluation and characterization of specimens suspected to contain pancreatic or bile duct cancer.