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49 High efficient circulating tumor cells (CTCs) mutation detection in cancer patients using targeted panel sequencing
  1. Robbie Huff,
  2. Dylan Dufek,
  3. Stephenie Jones,
  4. Janet Dickerson and
  5. Yipeng Wang
  1. BiFluidica, San Diego, CA, USA
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background Circulating tumor cells (CTCs) are rare cancer cells, shed from cancerous lesions into the bloodstream or other bodily fluids. Utilizing genomic characterization of CTCs presents a promising alternative to traditional tissue biopsies for profiling cancer mutations. However, the isolation of these scarce CTCs from whole blood poses significant challenges. Additionally, the limited amount of DNA available falls short of the requirements for next-generation sequencing (NGS) without the implementation of whole genome amplification (WGA). To address these obstacles, we have developed a fully automated solution for CTC enrichment, integrating WGA and targeted gene sequencing panel workflows.

Methods For this study, blood samples were collected from patients with breast, lung, and other solid tumors using BioFluidica blood collection tubes. The LiquidScan platform, employing microfluidic affinity selection with the EPCAM surface marker, was utilized to enrich CTCs. Cell eluates released from the microfluidic chips underwent further sorting to eliminate white blood cells, and the sorted cells were collected into multiple wells. To analyze the DNA, the cells underwent whole genome amplification using the Takara PicoPLEX single-cell kit and amplicon-based targeted sequencing utilizing the Illumina AmpiSeq Cancer Hotspot Panel v2 and the ArcherDx Solid Tumor Focus v2.

Results The whole blood samples were directly loaded onto microfluidic chips using a Hamilton robot for automated processing, minimizing manual intervention. The procedure did not require red blood cell lysis, and each sample took approximately 3 hours to process, with up to 8 samples simultaneously processed per instrument. The success rate of DNA amplifications exceeded 90%, and somatic mutations were detected in all patients.

Conclusions The LiquidScan platform offers a rapid and automated solution for CTC isolation, and we established workflows for detecting downstream somatic mutations. This study demonstrates the feasibility of utilizing CTCs for monitoring clinically relevant cancer mutations throughout a patient’s cancer therapy journey.

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See

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