title: Rapid, automated, parallel quantitative immunoassays using highly integrated microfluidics and AlphaLISA.
abstract: Immunoassays represent one of the most popular analytical methods for detection and quantification of biomolecules. However, conventional immunoassays such as ELISA and flow cytometry, even though providing high sensitivity and specificity and multiplexing capability, can be labor-intensive and prone to human error, making them unsuitable for standardized clinical diagnoses. Using a commercialized no-wash, homogeneous immunoassay technology ('AlphaLISA') in conjunction with integrated microfluidics, herein we developed a microfluidic immunoassay chip capable of rapid, automated, parallel immunoassays of microliter quantities of samples. Operation of the microfluidic immunoassay chip entailed rapid mixing and conjugation of AlphaLISA components with target analytes before quantitative imaging for analyte detections in up to eight samples simultaneously. Aspects such as fluid handling and operation, surface passivation, imaging uniformity, and detection sensitivity of the microfluidic immunoassay chip using AlphaLISA were investigated. The microfluidic immunoassay chip could detect one target analyte simultaneously for up to eight samples in 45 min with a limit of detection down to 10 pg mL(-1). The microfluidic immunoassay chip was further utilized for functional immunophenotyping to examine cytokine secretion from human immune cells stimulated ex vivo. Together, the microfluidic immunoassay chip provides a promising high-throughput, high-content platform for rapid, automated, parallel quantitative immunosensing applications.
authors.affiliation: Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Zeta Tak For
authors.initials: ZT
authors.lastname: Yu
authors.affiliation: Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Huijiao
authors.initials: H
authors.lastname: Guan
authors.affiliation: Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Mei Ki
authors.initials: MK
authors.lastname: Cheung
authors.affiliation: Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Walker M
authors.initials: WM
authors.lastname: McHugh
authors.affiliation: Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Timothy T
authors.initials: TT
authors.lastname: Cornell
authors.affiliation: Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Shanley
authors.affiliation: 1] Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA [2] Department of Electrical Engineering and Computer Science, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Katsuo
authors.initials: K
authors.lastname: Kurabayashi
authors.affiliation: 1] Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA [2] Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Jianping
authors.initials: J
authors.lastname: Fu
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry; D057205:Automation, Laboratory; D045744:Cell Line, Tumor; D004867:Equipment Design; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods*; D056656:Lab-On-A-Chip Devices; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects / Q000276:immunology; D057230:Limit of Detection; D008070:Lipopolysaccharides / Q000494:pharmacology; D046210:Microfluidic Analytical Techniques; D061251:Primary Cell Culture; D015203:Reproducibility of Results; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha