Separation and Analysis of Adherent and Non-Adherent Cancer Cells Using a Single-Cell Microarray Chip

Abstract

A new single-cell microarray chip was designed and developed to separate and analyze single adherent and non-adherent cancer cells. The single-cell microarray chip is made of polystyrene with over 60,000 microchambers of 10 different size patterns (31–40 µm upper diameter, 11–20 µm lower diameter). A drop of suspension of adherent carcinoma (NCI-H1650) and non-adherent leukocyte (CCRF-CEM) cells was placed onto the chip, and single-cell occupancy of NCI-H1650 and CCRF-CEM was determined to be 79% and 84%, respectively. This was achieved by controlling the chip design and surface treatment. Analysis of protein expression in single NCI-H1650 and CCRF-CEM cells was performed on the single-cell microarray chip by multi-antibody staining. Additionally, with this system, we retrieved positive single cells from the microchambers by a micromanipulator. Thus, this system demonstrates the potential for easy and accurate separation and analysis of various types of single cells.

Analysis of Single Nucleotide-Mutated Single-Cancer Cells Using the Combined Technologies of Single-Cell Microarray Chips and Peptide Nucleic Acid-DNA Probes

“…A further advantage of the single-cell microarray chip system is the ability to recover target single cells. Previously, we reported the recovery of single-cells by using a micromanipulator and a single-cell microarray chip [7]. Although we did not recover the T790M-mutated single-cancer cells from the microchambers in the present work, for further analysis it would also be easy to recover the mutated single-cancer cells by using a micromanipulator system in the future.…”

“…The surface of the single-cell microarray chip was hydrophilized with an oxygen plasma treatment using soft plasma-etching equipment (SEDE-PFA, Meiwafosis Co., Ltd., Tokyo, Japan), which made single-cell separation and analysis easier. The exposure time and electric current for plasma etching were described in previous studies [7].…”

“…In this study, we have developed a new detection system for single nucleotide-mutated cancer cells at the single-cell level using our original techniques combined with a single-cell microarray chip [7] and PNA-DNA probes [48]. After the T790M-mutated lung cancer cell line (NCI-H1975), as a model of anticancer drug-resistant cancer cells, was spiked into non-mutated cancer cells (A549), all cancer cells were separated into single cells using the single-cell microarray chip.…”

“…In particular, microchip technology, especially the microchip system for processing cells, called cell chips, could potentially be a powerful tool for the easy, rapid, accurate, and highly sensitive analysis of target single cells that exist within a large number of different cells. Many cell chips with types of microarray [ 4 , 5 , 6 , 7 , 8 , 9 ] and microfluidic [ 10 , 11 , 12 , 13 ] have been reported for single-cells analysis. In particular, cell microarray chips are useful for high-throughput screening and analysis for cells.…”

Time Sequential Single-Cell Patterning with High Efficiency and High Density

“…However, several malign mutations that cause maladies such as leukemia [ 1 ], liver cancer [ 2 ], and Alzheimer’s disease [ 3 ] usually start from a rare subpopulation of cells. The average-response-based analysis often fails to reveal some abnormal lesions because of the intrinsic drawbacks of this method, such as omission of “error” or “noise” signals [ 4 , 5 ]. The single-cell analysis focuses on the unique response of the subpopulation cells exposed to different stimuli [ 6 ].…”