title: High-Density Dielectrophoretic Microwell Array for Detection, Capture, and Single-Cell Analysis of Rare Tumor Cells in Peripheral Blood.
abstract: Development of a reliable platform and workflow to detect and capture a small number of mutation-bearing circulating tumor cells (CTCs) from a blood sample is necessary for the development of noninvasive cancer diagnosis. In this preclinical study, we aimed to develop a capture system for molecular characterization of single CTCs based on high-density dielectrophoretic microwell array technology. Spike-in experiments using lung cancer cell lines were conducted. The microwell array was used to capture spiked cancer cells, and captured single cells were subjected to whole genome amplification followed by sequencing. A high detection rate (70.2%-90.0%) and excellent linear performance (R2 = 0.8189-0.9999) were noted between the observed and expected numbers of tumor cells. The detection rate was markedly higher than that obtained using the CellSearch system in a blinded manner, suggesting the superior sensitivity of our system in detecting EpCAM- tumor cells. Isolation of single captured tumor cells, followed by detection of EGFR mutations, was achieved using Sanger sequencing. Using a microwell array, we established an efficient and convenient platform for the capture and characterization of single CTCs. The results of a proof-of-principle preclinical study indicated that this platform has potential for the molecular characterization of captured CTCs from patients.
authors.affiliation: Life Science Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Morimoto
authors.affiliation: Life Science Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Mogami
authors.affiliation: Third Department of Internal Medicine, Wakayama Medical University, Wakayama, Japan.
authors.forename: Masaru
authors.initials: M
authors.lastname: Watanabe
authors.affiliation: Life Science Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
authors.forename: Kazuki
authors.initials: K
authors.lastname: Iijima
authors.affiliation: Life Science Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Akiyama
authors.affiliation: Life Science Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Katayama
authors.affiliation: Life Science Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Futami
authors.affiliation: Third Department of Internal Medicine, Wakayama Medical University, Wakayama, Japan.
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Yamamoto
authors.affiliation: Shien-Lab, National Cancer Center Hospital, Tokyo, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sawada
authors.affiliation: Shien-Lab, National Cancer Center Hospital, Tokyo, Japan.
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Koizumi
authors.affiliation: Third Department of Internal Medicine, Wakayama Medical University, Wakayama, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Koh
mesh_terms: D045744:Cell Line, Tumor; D004586:Electrophoresis / Q000379:methods*; D006801:Humans; D009360:Neoplastic Cells, Circulating; D059010:Single-Cell Analysis