Related Experiment Video
Updated: Apr 16, 2026
Multiplex Cytokine Profiling of Stimulated Mouse Splenocytes Using a Cytometric Bead-based Immunoassay Platform
Published on: November 9, 2017
Multiplex serum cytokine immunoassay using nanoplasmonic biosensor microarrays
Pengyu Chen1, Meng Ting Chung1, Walker McHugh2
1†Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
Insights
This study introduces a novel localized surface plasmon resonance (LSPR) biosensor for rapid, multiplex cytokine analysis in small blood volumes. The high-sensitivity device enables precise immune status monitoring, crucial for critically ill patients and infants.
Area of Science:
- Biomedical Engineering
- Nanotechnology
- Immunology
Background:
- Precise immune status monitoring requires multiplex cytokine analysis from frequent blood samples.
- Current assays are impractical for clinical decision-making due to slow speed, low multiplexing, and large volume requirements.
- Infant immune monitoring is particularly challenging due to limited blood volume access.
Purpose of the Study:
- To develop a high-throughput, label-free, multiarrayed localized surface plasmon resonance (LSPR) optical biosensor for sensitive multiplex cytokine detection.
- To overcome the limited sensitivity of existing LSPR biosensing techniques for practical implementation in complex biological matrices.
- To enable rapid and sample-sparing immune monitoring for clinical applications, especially in vulnerable populations like infants.
Main Methods:
- Fabrication of a microfluidic device with 480 nanoplasmonic sensing spots using microfluidic patterning and gold nanorod (AuNR) antibody conjugation.
- Development of a high-throughput, label-free LSPR optical biosensing technique utilizing dark-field imaging to scan scattering light intensity.
- Demonstration of parallel multiplex immunoassays for six cytokines in a complex serum matrix using only 1 μL of sample.
Main Results:
- Achieved high-sensitivity quantitative cytokine measurements down to 5-20 pg/mL.
- Successfully demonstrated parallel multiplex detection of six cytokines in serum on a single chip.
- Completed whole on-chip assays, including sample loading, incubation, washing, and 10-fold replicated multianalyte detection, within 40 minutes.
- Validated the device's utility by monitoring inflammatory responses in infants post-cardiopulmonary bypass surgery.
Conclusions:
- The developed multiarrayed LSPR biosensor offers a rapid, sensitive, and sample-sparing solution for multiplex cytokine analysis.
- This technology overcomes previous sensitivity limitations, enabling practical implementation for immune status monitoring in complex samples.
- The biosensor is suitable for real-time monitoring of inflammatory responses, particularly in clinical settings involving infants and critically ill patients.
Abstract:
Precise monitoring of the rapidly changing immune status during the course of a disease requires multiplex analysis of cytokines from frequently sampled human blood. However, the current lack of rapid, multiplex, and low volume assays makes immune monitoring for clinical decision-making (e.g., critically ill patients) impractical. Without such assays, immune monitoring is even virtually impossible for infants and neonates with infectious diseases and/or immune mediated disorders as access to their blood in large quantities is prohibited. Localized surface plasmon resonance (LSPR)-based microfluidic optical biosensing is a promising approach to fill this technical gap as it could potentially permit real-time refractometric detection of biomolecular binding on a metallic nanoparticle surface and sensor miniaturization, both leading to rapid and sample-sparing analyte analysis. Despite this promise, practical implementation of such a microfluidic assay for cytokine biomarker detection in serum samples has not been established primarily due to the limited sensitivity of LSPR biosensing. Here, we developed a high-throughput, label-free, multiarrayed LSPR optical biosensor device with 480 nanoplasmonic sensing spots in microfluidic channel arrays and demonstrated parallel multiplex immunoassays of six cytokines in a complex serum matrix on a single device chip while overcoming technical limitations. The device was fabricated using easy-to-implement, one-step microfluidic patterning and antibody conjugation of gold nanorods (AuNRs). When scanning the scattering light intensity across the microarrays of AuNR ensembles with dark-field imaging optics, our LSPR biosensing technique allowed for high-sensitivity quantitative cytokine measurements at concentrations down to 5-20 pg/mL from a 1 μL serum sample. Using the nanoplasmonic biosensor microarray device, we demonstrated the ability to monitor the inflammatory responses of infants following cardiopulmonary bypass (CPB) surgery through tracking the time-course variations of their serum cytokines. The whole parallel on-chip assays, which involved the loading, incubation, and washing of samples and reagents, and 10-fold replicated multianalyte detection for each sample using the entire biosensor arrays, were completed within 40 min.
