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Enhanced chromosome extraction from cells using a pinched flow microfluidic device

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Abstract

Extraction and purification of intact chromosomes are critical sample preparation steps for transchromosomic research and other applications. The commonly used sample preparation methods lead to too few chromosomes with chromosome deactivation and degradation. In this paper, a “mild” chromosome extraction process that combines a chemical and mechanical lysis approach is introduced for the preparation of intact chromosomes that can readily be used for downstream processing. Metaphase cells are treated by chemical lysis buffer and pushed through a microfluidic pinched flow device. Cells are ruptured, and chromosomes are released by a combination of shear stress and chemical reagents. Chromosomes are released intact from the cell membrane into the solution. Simulations and experiments are performed to optimize the microfluidic device geometry and operation parameters. Cell rupture and chromosome release are found to be improved by the shear stress in the pinched flow device. Simulation results indicate that the maximum shear stress appears in the channel constriction region, and the narrow channel maintains constant shear stress. It is concluded that the constriction design, narrow channel width, and operation flow rate have a significate influence on chromosome release. Utilizing an optimized device, near-complete cell lysis is achieved and 4 times as many chromosomes are released (8% in control experiments to 25% in optimized pinched flow devices). Sample treatment time can also be reduced utilizing this combined chemical-mechanical chromosome release method.

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Acknowledgments

This work was supported by the University of Utah Flow Cytometry Facility in addition to the National Cancer Institute through Award Number 5P30CA042014-24.

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Authors and Affiliations

  1. State of Utah Center of Excellence for Biomedical Microfluidics, University of Utah, Salt Lake City, UT, USA

    Haidong Feng, Bruce Gale & Himanshu Sant

  2. The Capecchi Lab, Eccles Institute of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT, USA

    Matthew Hockin, Shuhua Zhang & Mario Capecchi

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  1. Haidong Feng
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  2. Matthew Hockin
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  3. Shuhua Zhang
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  4. Mario Capecchi
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Correspondence to Himanshu Sant.

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Feng, H., Hockin, M., Zhang, S. et al. Enhanced chromosome extraction from cells using a pinched flow microfluidic device. Biomed Microdevices 22, 25 (2020). https://doi.org/10.1007/s10544-020-0477-7

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  • DOI: https://doi.org/10.1007/s10544-020-0477-7

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