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Open AccessArticle

Absolute Copy Numbers of β-Actin Proteins Collected from 10,000 Single Cells

1
State Key Laboratory of Transducer Technology, Institute of Electronics, Chinese Academy of Sciences, Beijing 100190, China
2
University of Chinese Academy of Sciences, Beijing 100049, China
3
State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China
*
Authors to whom correspondence should be addressed.
Micromachines 2018, 9(5), 254; https://doi.org/10.3390/mi9050254
Received: 24 April 2018 / Revised: 14 May 2018 / Accepted: 14 May 2018 / Published: 22 May 2018
(This article belongs to the Special Issue Glassy Materials Based Microdevices)
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Abstract

Semi-quantitative studies have located varied expressions of β-actin proteins at the population level, questioning their roles as internal controls in western blots, while the absolute copy numbers of β-actins at the single-cell level are missing. In this study, a polymeric microfluidic flow cytometry was used for single-cell analysis, and the absolute copy numbers of single-cell β-actin proteins were quantified as 9.9 ± 4.6 × 105, 6.8 ± 4.0 × 105 and 11.0 ± 5.5 × 105 per cell for A549 (ncell = 14,754), Hep G2 (ncell = 36,949), and HeLa (ncell = 24,383), respectively. High coefficients of variation (~50%) and high quartile coefficients of dispersion (~30%) were located, indicating significant variations of β-actin proteins within the same cell type. Low p values (≪0.01) and high classification rates based on neural network (~70%) were quantified among A549, Hep G2 and HeLa cells, suggesting expression differences of β-actin proteins among three cell types. In summary, the results reported here indicate significant variations of β-actin proteins within the same cell type from cell to cell, and significant expression differences of β-actin proteins among different cell types, strongly questioning the properties of using β-actin proteins as internal controls in western blots. View Full-Text
Keywords: microfluidics; single-cell analysis; polymeric microfluidic flow cytometry; single-cell protein quantification microfluidics; single-cell analysis; polymeric microfluidic flow cytometry; single-cell protein quantification
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Fan, B.; Li, X.; Liu, L.; Chen, D.; Cao, S.; Men, D.; Wang, J.; Chen, J. Absolute Copy Numbers of β-Actin Proteins Collected from 10,000 Single Cells. Micromachines 2018, 9, 254.

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