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Related Experiment Video

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A Microfluidic Chip for the Versatile Chemical Analysis of Single Cells
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A Microfluidic Platform for Single Cell Fluorometric Granzyme B Profiling.

Jonathan C Briones1, Wilfred V Espulgar1, Shohei Koyama2

  • 1Graduate School of Engineering, Osaka University, Suita, Osaka, 565-0871, JAPAN.

Theranostics
|January 7, 2020
PubMed
Summary

This study developed a microfluidic platform to measure Granzyme B (GrB) activity in single cells, aiding immunotherapy efficacy prediction. The device successfully profiled GrB levels in model cells and patient samples, showing promise for personalized cancer treatment strategies.

Keywords:
granzyme b profilingimmunotherapymicrofluidicssingle cell

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Area of Science:

  • Biomedical Engineering
  • Immunology
  • Cancer Research

Background:

  • Granzyme B (GrB) is a key cytotoxic effector in cancer immunotherapy and a biomarker for predicting treatment efficacy.
  • Monitoring Granzyme B activity is crucial for tailoring immunotherapies, such as checkpoint inhibitors, to individual patients.
  • Current methods for Granzyme B assessment can be limiting for single-cell analysis and rapid clinical application.

Purpose of the Study:

  • To develop and validate a microfluidic platform for sensitive, single-cell Granzyme B activity measurement.
  • To demonstrate the platform's utility in distinguishing Granzyme B levels across different cell types.
  • To assess the potential of this platform for predicting patient response to immunotherapy.

Main Methods:

  • Fabrication of a microfluidic device utilizing hydrodynamic traps and pneumatic valving.
  • Fluorometric detection and quantification of single-cell Granzyme B activity using a commercial assay kit and a peptide substrate (Ac-IEPD-AFC).
  • Confocal and inverted microscopy for fluorescence observation and measurement in model cell lines (NK-92, Jurkat, THP1) and human PBMCs from healthy donors and lung cancer patients.

Main Results:

  • Differential Granzyme B activity was observed in model cell lines, with NK-92 cells showing higher activity than GrB-transduced Jurkat cells, and THP-1 cells exhibiting minimal activity.
  • A significant increase in Granzyme B expression was detected in PBMCs from an anti-PD-1 antibody-treated lung cancer patient compared to healthy donor PBMCs.
  • High Granzyme B activity in TCR+ Ig-G4+ PBMC cells indicated a positive response to PD-1 blockade therapy.

Conclusions:

  • The microfluidic platform successfully demonstrated the capability to measure single-cell Granzyme B enzymatic activity.
  • This platform shows promise as a tool for evaluating immunotherapy sensitivity.
  • The technology may aid in identifying specific T cell subsets crucial for anti-tumor responses and optimizing cancer treatment strategies.