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Screening of peptide specific to cholangiocarcinoma cancer cells using an integrated microfluidic system and phage display technology

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Abstract

Cholangiocarcinoma (CCA) is a cancer of the bile duct with high mortality rate and poor prognosis, owing to the difficulty in the early diagnosis and prognosis. The specific biomarkers or affinity reagents toward CCA cells could be great tools to assist in detection of CCA. However, screening of biomarkers/affinity reagents are generally labor-intensive, time-consuming and requiring large volume of samples and reagents. Therefore, we developed an integrated microfluidic system which could automatically perform selections of biomarkers and affinity reagents using phage display techniques. The experimental results showed that the selection of phage-displayed peptides bound to CCA cells was successfully demonstrated on the integrated microfluidic system using fewer reagents, samples and less time (5.25 h per biopanning round, and continuously performed for only 4 panning rounds). Three oligopeptides were screened, and their specificity and affinity toward CCA cells were characterized. Furthermore, comparing to conventional EpiEnrich beads for cancer cell capture, the screened CCA-specific peptides showed relatively low capture rate over control normal cells. It is envisioned that this microfluidic system may be a powerful tool for screening of biomarkers/affinity reagents in clinical diagnosis and target therapy for CCA.

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Abbreviations

bp:

Base pairs

CA19-9:

Cancer antigen 19-9

CCA:

Cholangiocarcinoma

CCD:

Charge-coupled-device

CEA:

Carcinoembryonic antigen

CNC:

Computer-numerical-control

DMEM:

Dulbecco’s modified eagle medium

ddH2O:

Double-stilled water

E. coli :

Escherichia coli

EphA5:

Ephrin type-A receptor 5

EphA2:

Ephrin type-A receptor 2

EpiEnrich:

Epithelial Enrich

FITC:

Fluorescein isothiocyanate

FGFR2:

Fibroblast growth factor receptor 2

FGFR3:

Fibroblast growth factor receptor 3

FBS:

Fetal bovine serum

IPTG/X-Gal:

Isopropyl β-D-1-thiogalactopyranoside/5-bromo-4-chloro-3-indolyl-d-galactoside

LB:

Luria–Bertani

NCKUH:

National Cheng Kung University Hospital

PBS:

Phosphate buffer saline

PCR:

Polymerase chain reaction

PDMS:

Polydimethylsiloxane

PMMA:

Polymethylmethacrylate

RPMI:

Roswell park memorial institute

SOC:

Super optimal broth with catabolite repression

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Acknowledgements

Authors would like to thank financial support from the National Health Research Institute in Taiwan (NHREI-EX104-10428EI) and Ministry of Science and Technology in Taiwan (MOST 105-2119-M-007-009).

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Authors and Affiliations

  1. Department of Power Mechanical Engineering, National Tsing Hua University, Hsinchu, 30013, Taiwan

    Ching-Wen Yu, Chien-Yu Fu, Lien-Yu Hung, Chih-Hung Wang & Gwo-Bin Lee

  2. Institute of Clinical Medicine, National Cheng Kung University Hospital, Tainan, 704, Taiwan

    Nai-Jung Chiang, Yu-Chun Wang & Yan-Shen Shan

  3. Institute of Biomedical Engineering, National Tsing Hua University, Hsinchu, 30013, Taiwan

    Gwo-Bin Lee

  4. Institute of NanoEngineering and Microsystems, National Tsing Hua University, Hsinchu, 30013, Taiwan

    Gwo-Bin Lee

  5. National Institute of Cancer Research, National Health Research Institutes, Tainan, 704, Taiwan

    Nai-Jung Chiang

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  1. Ching-Wen Yu
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Correspondence to Yan-Shen Shan or Gwo-Bin Lee.

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Yu, CW., Fu, CY., Hung, LY. et al. Screening of peptide specific to cholangiocarcinoma cancer cells using an integrated microfluidic system and phage display technology. Microfluid Nanofluid 21, 145 (2017). https://doi.org/10.1007/s10404-017-1983-7

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