Microfluidic immunocapture of circulating pancreatic cells using parallel EpCAM and MUC1 capture: Characterization, optimization and downstream analysis

Fredrik I. Thege, Timothy B. Lannin, Trisha N. Saha, Shannon Tsai, Michael L. Kochman, Michael A Hollingsworth, Andrew D. Rhim, Brian J. Kirby

Research output: Contribution to journalArticle

64 Citations (Scopus)

Abstract

We have developed and optimized a microfluidic device platform for the capture and analysis of circulating pancreatic cells (CPCs) and pancreatic circulating tumor cells (CTCs). Our platform uses parallel anti-EpCAM and cancer-specific mucin 1 (MUC1) immunocapture in a silicon microdevice. Using a combination of anti-EpCAM and anti-MUC1 capture in a single device, we are able to achieve efficient capture while extending immunocapture beyond single marker recognition. We also have detected a known oncogenic KRAS mutation in cells spiked in whole blood using immunocapture, RNA extraction, RT-PCR and Sanger sequencing. To allow for downstream single-cell genetic analysis, intact nuclei were released from captured cells by using targeted membrane lysis. We have developed a staining protocol for clinical samples, including standard CTC markers; DAPI, cytokeratin (CK) and CD45, and a novel marker of carcinogenesis in CPCs, mucin 4 (MUC4). We have also demonstrated a semi-automated approach to image analysis and CPC identification, suitable for clinical hypothesis generation. Initial results from immunocapture of a clinical pancreatic cancer patient sample show that parallel capture may capture more of the heterogeneity of the CPC population. With this platform, we aim to develop a diagnostic biomarker for early pancreatic carcinogenesis and patient risk stratification.

Original languageEnglish (US)
Pages (from-to)1775-1784
Number of pages10
JournalLab on a Chip
Volume14
Issue number10
DOIs
StatePublished - Jan 1 2014

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Mucin-1
Microfluidics
Tumors
Mucin-4
Cells
Biomarkers
Silicon
Keratins
RNA
Image analysis
Circulating Neoplastic Cells
Blood
Lab-On-A-Chip Devices
Membranes
Carcinogenesis
Single-Cell Analysis
Tumor Biomarkers
Clinical Protocols
Pancreatic Neoplasms
Epithelial Cell Adhesion Molecule

ASJC Scopus subject areas

  • Bioengineering
  • Biochemistry
  • Chemistry(all)
  • Biomedical Engineering

Cite this

Microfluidic immunocapture of circulating pancreatic cells using parallel EpCAM and MUC1 capture : Characterization, optimization and downstream analysis. / Thege, Fredrik I.; Lannin, Timothy B.; Saha, Trisha N.; Tsai, Shannon; Kochman, Michael L.; Hollingsworth, Michael A; Rhim, Andrew D.; Kirby, Brian J.

In: Lab on a Chip, Vol. 14, No. 10, 01.01.2014, p. 1775-1784.

Research output: Contribution to journalArticle

Thege, Fredrik I. ; Lannin, Timothy B. ; Saha, Trisha N. ; Tsai, Shannon ; Kochman, Michael L. ; Hollingsworth, Michael A ; Rhim, Andrew D. ; Kirby, Brian J. / Microfluidic immunocapture of circulating pancreatic cells using parallel EpCAM and MUC1 capture : Characterization, optimization and downstream analysis. In: Lab on a Chip. 2014 ; Vol. 14, No. 10. pp. 1775-1784.
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