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Microfilter-Based Capture and Release of Viable Circulating Tumor Cells

  • Siddarth Rawal
  • Zheng Ao
  • Ram H. Datar
  • Ashutosh Agarwal
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1634)

Abstract

Microfilters with slot-pore geometry can be used for size-based capture of circulating tumor cells (CTC) from the blood of cancer patients. The slot pore geometry reduces the shear stress that the cells would typically experience during filtration process and allows the cells to remain viable. The microfilter provides a platform capable of high CTC capture efficiency; however, the release of these cells from the filter following capture is nontrivial, possibly due to the strong nonspecific electrostatic adhesion of CTC to the microfilter surface. Techniques such as reverse flow or cell scraping result in recovery of only a small percentage of captured cells. We describe, in detail, a protocol for novel application of thermo-responsive polymer poly(N-iso-propylacrylamide) (PIPAAm) to release viable CTCs from microfilters with slot pores. Following fabrication of the microfilter, a coating of PIPAAm is applied to the surface to exploit its thermoresponsive interfacial properties to release the cells. Typically, cancer patient’s blood is filtered at room temperature (below 32 °C) when PIPAAm is hydrophilic. Thereafter, the filter is placed in either culture medium or a buffer maintained at 37 °C, which renders PIPAAm hydrophobic, allowing subsequent release of the electrostatically bound cells with high efficiency. Using this method, viable CTC captured directly from cancer patients’ blood can be subjected to downstream off-chip culture, analyses, and characterization.

Key words

Circulating tumor cell CTC culture Viable CTC Microfilter Live cell capture Precision medicine Personalized medicine Translational medicine Liquid biopsy 

Notes

Acknowledgments

We thank all the patients who have donated blood samples to support this work. We thank Drs. Guiseppe Giaconne, Ritesh Parajuli, and Marc E. Lippman for their assistance in clinical sample acquirement, and Drs. Carmen Gomez, Ralf Landgraf, Stephan Züchner, Toumy Guettouche, Diana Lopez for their insightful discussions. Zheng Ao thanks partial support and assistance from the Sheila and David Fuente Graduate Program in Cancer Biology, Sylvester Comprehensive Cancer Center.

Disclosures

Part of this work has been protected under U.S. Patent Application No. 15/269,574. Ram Datar and Siddarth Rawal are shareholders of Circulogix Inc. that commercially produces the filters and filter cassettes/cartridges used in this chapter.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Siddarth Rawal
    • 1
    • 2
  • Zheng Ao
    • 2
    • 3
  • Ram H. Datar
    • 2
  • Ashutosh Agarwal
    • 1
    • 2
  1. 1.Department of Biomedical EngineeringUniversity of MiamiCoral GablesUSA
  2. 2.Department of PathologyUniversity of MiamiCoral GablesUSA
  3. 3.SRI InternationalMenlo ParkUSA

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