title: Multiplex serum cytokine immunoassay using nanoplasmonic biosensor microarrays.
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.
authors: †Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Pengyu
authors: P
authors: Chen
authors: †Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Meng Ting
authors: MT
authors: Chung
authors: ‡Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Walker
authors: W
authors: McHugh
authors: †Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Robert
authors: R
authors: Nidetz
authors: †Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Yuwei
authors: Y
authors: Li
authors: †Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.|§Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Jianping
authors: J
authors: Fu
authors: ‡Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Timothy T
authors: TT
authors: Cornell
authors: ‡Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Thomas P
authors: TP
authors: Shanley
authors: †Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.|⊥Department of Electrical Engineering and Computer Science, University of Michigan, Ann Arbor, Michigan 48109, United States.
authors: Katsuo
authors: K
authors: Kurabayashi
mesh_terms: D002315:Cardiopulmonary Bypass; D016207:Cytokines / Q000097:blood*; D006801:Humans; D007109:Immunity; D007118:Immunoassay / Q000379:methods*; D007223:Infant; D008991:Monitoring, Physiologic; D036103:Nanotechnology / Q000379:methods*; D020349:Surface Plasmon Resonance / Q000379:methods*
chemical_list: D016207:Cytokines
keywords: localized surface plasmon resonance (LSPR); multiplexed immunoassay; nanoplasmonic sensing; optofluidics; serum cytokines