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Direct Write Protein Patterns for Multiplexed Cytokine Detection from Live Cells Using Electron Beam Lithography.

Uland Y Lau1, Sina S Saxer2, Juneyoung Lee2

  • 1Department of Bioengineering, University of California, Los Angeles , 410 Westwood Plaza, Los Angeles, California 90095, United States.

ACS Nano
|December 19, 2015
PubMed
Summary

This study presents a novel method for precisely immobilizing antibodies on silicon surfaces using electron beam lithography. This technique enables multiplexed cytokine detection for improved disease diagnostics and biosensing applications.

Keywords:
antibody patterningbiosensordark-field microscopyelectron beam lithographylocalized surface plasmon resonancetrehalose glycopolymer

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Area of Science:

  • Biotechnology
  • Nanotechnology
  • Immunology

Background:

  • Simultaneous detection of multiple extracellular signaling molecules is crucial for disease diagnostics.
  • Miniaturization of assays for parallel, fast, low-volume detection remains a challenge.
  • Precise antibody positioning at the micro- and nanoscale is key for improving biosensors and diagnostics.

Purpose of the Study:

  • To develop a multiplexed cytokine immunoassay with micro- and nanoscale precision.
  • To demonstrate direct antibody patterning on silicon substrates for enhanced biosensing.
  • To validate the retention of antibody binding properties after direct patterning.

Main Methods:

  • Utilized electron beam lithography and a trehalose glycopolymer resist for direct antibody writing.
  • Patterned antibodies against interleukin-6 (IL-6) and tumor necrosis factor alpha (TNFα).
  • Employed a sandwich immunoassay with gold nanoparticles and silver enhancement for detection via localized surface plasmon resonance (LSPR) and dark-field microscopy.

Main Results:

  • Successfully demonstrated direct, high-precision immobilization of anti-IL-6 and anti-TNFα antibodies on silicon.
  • Confirmed retention of specific antibody binding properties by capturing cytokines from stimulated macrophages.
  • Achieved successful multiplexed detection of both IL-6 and TNFα on a single chip with high specificity.

Conclusions:

  • Direct fabrication of capture antibody patterns using electron beam lithography offers micro- and nanoscale precision for biosensing.
  • This method enables multiplexed cytokine detection, advancing disease profiling and diagnostics.
  • The developed technique is suitable for applications requiring sensitive and specific detection of multiple biomarkers in relevant biological conditions.