title: Development of on-chip multi-imaging flow cytometry for identification of imaging biomarkers of clustered circulating tumor cells.
abstract: An on-chip multi-imaging flow cytometry system has been developed to obtain morphometric parameters of cell clusters such as cell number, perimeter, total cross-sectional area, number of nuclei and size of clusters as "imaging biomarkers", with simultaneous acquisition and analysis of both bright-field (BF) and fluorescent (FL) images at 200 frames per second (fps); by using this system, we examined the effectiveness of using imaging biomarkers for the identification of clustered circulating tumor cells (CTCs). Sample blood of rats in which a prostate cancer cell line (MAT-LyLu) had been pre-implanted was applied to a microchannel on a disposable microchip after staining the nuclei using fluorescent dye for their visualization, and the acquired images were measured and compared with those of healthy rats. In terms of the results, clustered cells having (1) cell area larger than 200 µm2 and (2) nucleus area larger than 90 µm2 were specifically observed in cancer cell-implanted blood, but were not observed in healthy rats. In addition, (3) clusters having more than 3 nuclei were specific for cancer-implanted blood and (4) a ratio between the actual perimeter and the perimeter calculated from the obtained area, which reflects a shape distorted from ideal roundness, of less than 0.90 was specific for all clusters having more than 3 nuclei and was also specific for cancer-implanted blood. The collected clusters larger than 300 µm2 were examined by quantitative gene copy number assay, and were identified as being CTCs. These results indicate the usefulness of the imaging biomarkers for characterizing clusters, and all of the four examined imaging biomarkers-cluster area, nuclei area, nuclei number, and ratio of perimeter-can identify clustered CTCs in blood with the same level of preciseness using multi-imaging cytometry.
authors.affiliation: Kanagawa Academy of Science and Technology, Takatsu, Kawasaki, Japan.
authors.forename: Hyonchol
authors.initials: H
authors.lastname: Kim
authors.affiliation: Kanagawa Academy of Science and Technology, Takatsu, Kawasaki, Japan; Department of Biomedical Information, Division of Biosystems, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Chiyoda, Tokyo, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Terazono
authors.affiliation: Molecular Pathology and Genetics Division, Kanagawa Cancer Center Research Institute, Asahi-ku, Yokohama, Japan.
authors.forename: Yoshiyasu
authors.initials: Y
authors.lastname: Nakamura
authors.affiliation: Department of Genome Biology, School of Medicine, Kinki University, Osaka-Sayama, Osaka, Japan.
authors.forename: Kazuko
authors.initials: K
authors.lastname: Sakai
authors.affiliation: Kanagawa Academy of Science and Technology, Takatsu, Kawasaki, Japan.
authors.forename: Akihiro
authors.initials: A
authors.lastname: Hattori
authors.affiliation: Kanagawa Academy of Science and Technology, Takatsu, Kawasaki, Japan.
authors.forename: Masao
authors.initials: M
authors.lastname: Odaka
authors.affiliation: Kanagawa Academy of Science and Technology, Takatsu, Kawasaki, Japan.
authors.forename: Mathias
authors.initials: M
authors.lastname: Girault
authors.affiliation: Department of Genome Biology, School of Medicine, Kinki University, Osaka-Sayama, Osaka, Japan.
authors.forename: Tokuzo
authors.initials: T
authors.lastname: Arao
authors.affiliation: Department of Genome Biology, School of Medicine, Kinki University, Osaka-Sayama, Osaka, Japan.
authors.forename: Kazuto
authors.initials: K
authors.lastname: Nishio
authors.affiliation: Molecular Pathology and Genetics Division, Kanagawa Cancer Center Research Institute, Asahi-ku, Yokohama, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Miyagi
authors.affiliation: Kanagawa Academy of Science and Technology, Takatsu, Kawasaki, Japan; Department of Biomedical Information, Division of Biosystems, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Chiyoda, Tokyo, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Yasuda
mesh_terms: D000818:Animals; D014408:Biomarkers, Tumor / Q000032:analysis*; D045744:Cell Line, Tumor; D003952:Diagnostic Imaging; D005434:Flow Cytometry / Q000379:methods*; D008297:Male; D009360:Neoplastic Cells, Circulating / Q000473:pathology*; D011471:Prostatic Neoplasms / Q000175:diagnosis* / Q000473:pathology; D051381:Rats
chemical_list: D014408:Biomarkers, Tumor