Tumor Cell Detection among Leukocytes by Microchip
Yamamura, Shohei National Institute of Advanced Industrial Science and Technology
Yatsushiro, Shouki National Institute of Advanced Industrial Science and Technology
Yamaguchi, Yuka National Institute of Advanced Industrial Science and Technology Tokushima University Educator and Researcher Directory
Abe, Kaori National Institute of Advanced Industrial Science and Technology KAKEN Search Researchers
Shinohara, Yasuo University of Tokushima Tokushima University Educator and Researcher Directory KAKEN Search Researchers
Tamiya, Eiichi Osaka University
Baba, Yoshinobu National Institute of Advanced Industrial Science and Technology|Nagoya University
Kataoka, Masatoshi National Institute of Advanced Industrial Science and Technology
Background: Accurate detection and analysis of circulating tumor cells plays an important role in the diagnosis and treatment of metastatic cancer treatment.
Methods and Findings: A cell microarray chip was used to detect spiked carcinoma cells among leukocytes. The chip, with 20,944 microchambers (105 µm width and 50 µm depth), was made from polystyrene; and the formation of monolayers of leukocytes in the microchambers was observed. Cultured human T lymphoblastoid leukemia (CCRF-CEM) cells were used to examine the potential of the cell microarray chip for the detection of spiked carcinoma cells. A T lymphoblastoid leukemia suspension was dispersed on the chip surface, followed by 15 min standing to allow the leukocytes to settle down into the microchambers. Approximately 29 leukocytes were found in each microchamber when about 600,000 leukocytes in total were dispersed onto a cell microarray chip. Similarly, when leukocytes isolated from human whole blood were used, approximately 89 leukocytes entered each microchamber when about 1,800,000 leukocytes in total were placed onto the cell microarray chip. After washing the chip surface, PE-labeled anti-cytokeratin monoclonal antibody and APC-labeled anti-CD326 (EpCAM) monoclonal antibody solution were dispersed onto the chip surface and allowed to react for 15 min; and then a microarray scanner was employed to detect any fluorescence-positive cells within 20 min. In the experiments using spiked carcinoma cells (NCI-H1650, 0.01 to 0.0001%), accurate detection of carcinoma cells was achieved with PE-labeled anti-cytokeratin monoclonal antibody. Furthermore, verification of carcinoma cells in the microchambers was performed by double staining with the above monoclonal antibodies.
Conclusion: The potential application of the cell microarray chip for the detection of CTCs was shown, thus demonstrating accurate detection by double staining for cytokeratin and EpCAM at the single carcinoma cell level.
© 2012 Yamamura et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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pone_7_3_e32370.pdf 2.74 MB
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