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Isolation and Flow Cytometric Analysis of Immune Cells from the Ischemic Mouse Brain
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Parallel affinity-based isolation of leukocyte subsets using microfluidics: application for stroke diagnosis.

Swathi R Pullagurla1, Małgorzata A Witek, Joshua M Jackson

  • 1Department of Chemistry, Louisiana State University , Baton Rouge, Louisiana 70803, United States.

Analytical Chemistry
|March 22, 2014
PubMed
Summary

This study introduces a polymer microfluidic device for simultaneous isolation of multiple cell types, enabling gene expression profiling. The device achieves high purity and recovery, crucial for disease diagnosis like stroke.

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Area of Science:

  • Biomedical Engineering
  • Cell Biology
  • Molecular Diagnostics

Background:

  • Simultaneous isolation of specific cell populations from complex biological samples is challenging.
  • Accurate gene expression profiling requires high-purity cell populations.
  • Current methods for cell isolation can be time-consuming and have limited throughput.

Purpose of the Study:

  • To design and evaluate a polymer microfluidic device for simultaneous affinity selection of multiple cell types.
  • To assess the purity, recovery, and processing time of the microfluidic cell isolation.
  • To demonstrate the utility of isolated cells for downstream gene expression analysis.

Main Methods:

  • Development of a polymer microfluidic device with antibody-modified selection beds.
  • Evaluation of different device geometries (bifurcated, Z-configured) for cell selection efficiency.
  • Simultaneous isolation of CD4+ T-cells and neutrophils from whole blood using affinity-based capture.

Main Results:

  • Simultaneous isolation of CD4+ T-cells and neutrophils with >90% purity in approximately 3 minutes.
  • Sufficient cell recovery from small blood volumes (50 μL) for downstream molecular analysis.
  • Successful gene expression profiling (RT-PCR) of isolated T-cells and neutrophils, identifying key genes.

Conclusions:

  • The developed microfluidic device enables efficient and simultaneous isolation of multiple cell types with high purity.
  • This technology facilitates gene expression profiling for diagnostic applications, such as stroke-related gene analysis.
  • The platform is adaptable for isolating and analyzing various cell subsets from biological samples.