title: Single Cell Analysis of Leukocyte Protease Activity Using Integrated Continuous-Flow Microfluidics.
abstract: Leukocytes are the essential cells of the immune system that protect the human body against bacteria, viruses, and other foreign invaders. Secretory products of individual leukocytes, such as matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinase (ADAMs), are critical for regulating the inflammatory response and mediating host defense. Conventional single cell analytical methods, such as flow cytometry for cellular surface biomarker studies, are insufficient for performing functional assays of the protease activity of individual leukocytes. Here, an integrated continuous-flow microfluidic assay is developed to effectively detect secretory protease activity of individual viable leukocytes. Leukocytes in blood are first washed on-chip with defined buffer to remove background activity, followed by encapsulating individual leukocytes with protease sensors in water-in-oil droplets and incubating for 1 h to measure protease secretion. With this design, single leukocyte protease profiles under naive and phorbol 12-myristate 13-acetate (PMA)-stimulated conditions are reliably measured. It is found that PMA treatment not only elevates the average protease activity level but also reduces the cellular heterogeneity in protease secretion, which is important in understanding immune capability and the disease condition of individual patients.
authors: Department of Biomedical Engineering, National University of Singapore , Singapore 119077.|Singapore-MIT Alliance for Research and Technology , Singapore 138602.
authors: Tengyang
authors: T
authors: Jing
authors: Department of Biomedical Engineering, National University of Singapore , Singapore 119077.
authors: Zhangxing
authors: Z
authors: Lai
authors: Lidan
authors: L
authors: Wu
authors: Singapore-MIT Alliance for Research and Technology , Singapore 138602.
authors: Jongyoon
authors: J
authors: Han
authors: Department of Biomedical Engineering, National University of Singapore , Singapore 119077.|Singapore-MIT Alliance for Research and Technology , Singapore 138602.|Mechanobiology Institute , Singapore 117411.
authors: Chwee Teck
authors: CT
authors: Lim
authors: Department of Biomedical Engineering, National University of Singapore , Singapore 119077.|Singapore-MIT Alliance for Research and Technology , Singapore 138602.|Singapore Institute for Neurotechnology (SINAPSE) , Singapore 117456.
authors: Chia-Hung
authors: CH
authors: Chen
mesh_terms: D002470:Cell Survival; D004058:Diffusion; D004867:Equipment Design; D006801:Humans; D007962:Leukocytes / Q000166:cytology* / Q000201:enzymology*; D046210:Microfluidic Analytical Techniques; D010447:Peptide Hydrolases / Q000378:metabolism*; D059010:Single-Cell Analysis; D012984:Software
chemical_list: D010447:Peptide Hydrolases