title: High-Throughput Microdissection for Next-Generation Sequencing.
abstract: Precision medicine promises to enhance patient treatment through the use of emerging molecular technologies, including genomics, transcriptomics, and proteomics. However, current tools in surgical pathology lack the capability to efficiently isolate specific cell populations in complex tissues/tumors, which can confound molecular results. Expression microdissection (xMD) is an immuno-based cell/subcellular isolation tool that procures targets of interest from a cytological or histological specimen. In this study, we demonstrate the accuracy and precision of xMD by rapidly isolating immunostained targets, including cytokeratin AE1/AE3, p53, and estrogen receptor (ER) positive cells and nuclei from tissue sections. Other targets procured included green fluorescent protein (GFP) expressing fibroblasts, in situ hybridization positive Epstein-Barr virus nuclei, and silver stained fungi. In order to assess the effect on molecular data, xMD was utilized to isolate specific targets from a mixed population of cells where the targets constituted only 5% of the sample. Target enrichment from this admixed cell population prior to next-generation sequencing (NGS) produced a minimum 13-fold increase in mutation allele frequency detection. These data suggest a role for xMD in a wide range of molecular pathology studies, as well as in the clinical workflow for samples where tumor cell enrichment is needed, or for those with a relative paucity of target cells.
authors.affiliation: Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Avi Z
authors.initials: AZ
authors.lastname: Rosenberg
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Michael D
authors.initials: MD
authors.lastname: Armani
authors.affiliation: Cytopathology Section, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Fetsch
authors.affiliation: Molecular Diagnostics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Liqiang
authors.initials: L
authors.lastname: Xi
authors.affiliation: Molecular Diagnostics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Tina Thu
authors.initials: TT
authors.lastname: Pham
authors.affiliation: Molecular Diagnostics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Mark
authors.initials: M
authors.lastname: Raffeld
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.|Department of Mechanical Engineering, Johns Hopkins University, Baltimore, Maryland, United States of America.
authors.forename: Yun
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Neil
authors.initials: N
authors.lastname: O'Flaherty
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Stussman
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Adele R
authors.initials: AR
authors.lastname: Blackler
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Du
authors.affiliation: Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Jeffrey C
authors.initials: JC
authors.lastname: Hanson
authors.affiliation: Cytopathology Section, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Roth
authors.affiliation: Cytopathology Section, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Armando C
authors.initials: AC
authors.lastname: Filie
authors.affiliation: Department of Pathology, University of Michigan, Ann Arbor, Michigan, United States of America.
authors.forename: Michael H
authors.initials: MH
authors.lastname: Roh
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.|Avoneaux Medical Institute, Oxford, Maryland, United States of America.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Emmert-Buck
authors.affiliation: Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Jason D
authors.initials: JD
authors.lastname: Hipp
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.|Alvin & Lois Lapidus Cancer Institute, Sinai Hospital, Baltimore, Maryland, United States of America.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Tangrea
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D051379:Mice; D042282:Microdissection / Q000379:methods*; D041681:NIH 3T3 Cells; D013194:Staining and Labeling