title: Multiplexed femtomolar quantitation of human cytokines in a fluoropolymer microcapillary film.
abstract: Sensitive quantitation of multiple cytokines can provide important diagnostic information during infection, inflammation and immunopathology. In this study sensitive immunoassay detection of human cytokines IL-1β, IL-6, IL-12p70 and TNFα is shown for singleplex and multiplex formats using a novel miniaturized ELISA platform. The platform uses a disposable plastic multi-syringe aspirator (MSA) integrating 8 disposable fluoropolymer microfluidic test strips, each containing an array of ten 200 μm mean i.d. microcapillaries coated with a set of monoclonal antibodies. Each MSA device thus performs 10 tests on 8 samples, delivering 80 measurements. Unprecedented levels of sensitivity were obtained with the novel fluoropolymer microfluidic material and simple colorimetric detection in a flatbed scanner. The limit of detection for singleplex detection ranged from 2.0 to 15.0 pg ml(-1), i.e. 35 and 713 femtomolar for singleplex cytokine detection, and the intra- and inter-assay coefficient of variation (CV) remained within 10%. In addition, a triplex immunoassay was developed for measuring IL-1β, IL-12p70 and TNFα simultaneously from a given sample in the pg ml(-1) range. These assays permit high sensitivity measurement with rapid <15 min assay or detection from undiluted blood serum. The portability, speed and low-cost of this system are highly suited to point-of-care testing and field diagnostics applications.
authors.affiliation: Capillary Film Technology Ltd, Billingshurst, West Sussex RH14 9SJ, UK.
authors.forename: Ana P
authors.initials: AP
authors.lastname: Castanheira
authors.forename: Ana I
authors.initials: AI
authors.lastname: Barbosa
authors.forename: Alexander D
authors.initials: AD
authors.lastname: Edwards
authors.forename: Nuno M
authors.initials: NM
authors.lastname: Reis
mesh_terms: D001774:Blood Chemical Analysis / Q000379:methods*; D003124:Colorimetry; D016207:Cytokines / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay; D005461:Fluorine / Q000737:chemistry; D006801:Humans; D057230:Limit of Detection; D011108:Polymers / Q000737:chemistry*; D013499:Surface Properties; D013997:Time Factors
chemical_list: D016207:Cytokines; D011108:Polymers; D005461:Fluorine