title: Digital Sorting of Pure Cell Populations Enables Unambiguous Genetic Analysis of Heterogeneous Formalin-Fixed Paraffin-Embedded Tumors by Next Generation Sequencing.
abstract: Precision medicine in oncology requires an accurate characterization of a tumor molecular profile for patient stratification. Though targeted deep sequencing is an effective tool to detect the presence of somatic sequence variants, a significant number of patient specimens do not meet the requirements needed for routine clinical application. Analysis is hindered by contamination of normal cells and inherent tumor heterogeneity, compounded with challenges of dealing with minute amounts of tissue and DNA damages common in formalin-fixed paraffin-embedded (FFPE) specimens. Here we present an innovative workflow using DEPArray™ system, a microchip-based digital sorter to achieve 100%-pure, homogenous subpopulations of cells from FFPE samples. Cells are distinguished by fluorescently labeled antibodies and DNA content. The ability to address tumor heterogeneity enables unambiguous determination of true-positive sequence variants, loss-of-heterozygosity as well as copy number variants. The proposed strategy overcomes the inherent trade-offs made between sensitivity and specificity in detecting genetic variants from a mixed population, thus rescuing for analysis even the smaller clinical samples with low tumor cellularity.
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Chiara
authors: C
authors: Bolognesi
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Claudio
authors: C
authors: Forcato
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Genny
authors: G
authors: Buson
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Francesca
authors: F
authors: Fontana
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Chiara
authors: C
authors: Mangano
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Anna
authors: A
authors: Doffini
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Valeria
authors: V
authors: Sero
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Rossana
authors: R
authors: Lanzellotto
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Giulio
authors: G
authors: Signorini
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Alex
authors: A
authors: Calanca
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Maximilian
authors: M
authors: Sergio
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Rita
authors: R
authors: Romano
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Stefano
authors: S
authors: Gianni
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Gianni
authors: G
authors: Medoro
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Giuseppe
authors: G
authors: Giorgini
authors: Department of Pathology, Leiden University Medical Center, Leiden, Netherlands.
authors: Hans
authors: H
authors: Morreau
authors: European Institute of Oncology, Milan, Italy.
authors: Massimo
authors: M
authors: Barberis
authors: Department of Pathology, Leiden University Medical Center, Leiden, Netherlands.
authors: Willem E
authors: WE
authors: Corver
authors: Silicon Biosystems S.p.A, Bologna, Italy.
authors: Nicolò
authors: N
authors: Manaresi
mesh_terms: D002469:Cell Separation / Q000295:instrumentation / Q000379:methods*; D056915:DNA Copy Number Variations; D005404:Fixatives; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D005557:Formaldehyde; D014644:Genetic Variation; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D046228:Microarray Analysis / Q000295:instrumentation / Q000379:methods*; D009154:Mutation; D009369:Neoplasms / Q000175:diagnosis* / Q000235:genetics / Q000473:pathology; D016612:Paraffin Embedding; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D016707:Tissue Fixation
chemical_list: D005404:Fixatives; D005557:Formaldehyde