Journal
BIOSENSORS & BIOELECTRONICS
Volume 139, Issue -, Pages -Publisher
ELSEVIER ADVANCED TECHNOLOGY
DOI: 10.1016/j.bios.2019.111326
Keywords
Circulating tumor cells (CTCs); Micro-well array; Lung cancer; EGFR point mutation; Single-cell genetic analysis; Heterogeneity
Categories
Funding
- National Key Research and Development Program of China [2017YFA0205303, 20172X10302201-005-006]
- National Natural Science Foundation of China [61871243, 61671284, 61801465]
- Science and Technology Commission of Shanghai Municipality [17JC1401001]
- Research project of scientific research equipment of CAS [YJKYYQ20170043]
- STS program [KFJ-STS-ZDTP-061]
- science and technology support plan of the Shanghai Science and Technology Committee [17441904300]
- Zhejiang provincial natural science foundation [LQ19F010004]
- China Postdoctoral Science Foundation [2018M642384]
- K.C.Wong Magna Fund in Ningbo University
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In view of their critical function in metastasis, characterization of single circulating tumor cells (CTCs) can provide important clinical information to monitor tumor progression and guide personal therapy. Single-cell genetic analysis methods based on microfluidics have some inherent shortcomings such as complicated operation, low throughput, and expensive equipment requirements. To overcome these barriers, we developed a simple and open micro-well array containing 26,208 units for either nuclear acids or single-cell genetic analysis. Through modification of the polydimethylsiloxane surface and optimization of chip packaging, we addressed protein adsorption and solution evaporation for PCR amplification on a chip. In the detection of epidermal growth factor receptor (EGFR) exon gene 21, this micro-well array demonstrated good linear correlation at a DNA concentration from 1 x 10(1) to 1 x 10(5) copies/mu L (R-2 = 0.9877). We then successfully integrated cell capture, lysis, PCR amplification, and signal read-out on the micro-well array, enabling the rapid and simple genetic analysis of single cells. This device was used to detect duplex EGFR mutation genes of lung cancer cell lines (H1975 and A549 cells) and normal leukocytes, demonstrating the ability to perform high-throughput, massively parallel duplex gene analysis at the single-cell level. Different types of point mutations (EGFR-L858R mutation or EGFR-T790M mutation) were detected in single H1975 cells, further validating the significance of single-cell level gene detection. In addition, this method showed a good performance in the heterogeneity detection of individual CTCs from lung cancer patients, required for micro-invasive cancer monitoring and treatment selection.
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