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An antibody, specific to fibrinogen, has been covalently attached to graphene and deposited onto screen printed electrodes using a chitosan hydrogel binder to prepare an inexpensive electrochemical fibrinogen biosensor. Fourier Transform Infrared (FT-IR) spectroscopy has been utilized to confirm the presence of the antibody on the graphene scaffold. Electrochemical Impedance Spectroscopy (EIS) has been utilized to demonstrate that the biosensor responds in a selective manner to fibrinogen in aqueous media even in the presence of plasminogen, a potentially interfering molecule in the coagulopathy cascade. Furthermore, the biosensor was shown to reliably sense fibrinogen in the presence of high background serum albumin levels. Finally, we demonstrated detection of clinically relevant fibrinogen concentrations (938-44,542μg/dL) from human serum and human whole blood samples using this biosensor. This biosensor can potentially be used in a point-of-care device to detect the onset of coagulopathy and monitor response following therapeutic intervention in trauma patients. Thus this biosensor may improve the clinical management of patients with trauma-induced coagulopathy.

[This corrects the article DOI: 10.1371/journal.pone.0186782.].

Demand for highly sensitive, robust diagnostics and environmental monitoring methods has led to extensive research in improving reporter technologies. Inorganic phosphorescent materials exhibiting persistent luminescence are commonly found in electroluminescent displays and glowing paints but are not widely used as reporters in diagnostic assays. Persistent luminescence nanoparticles (PLNPs) offer advantages over conventional photoluminescent probes, including the potential for enhanced sensitivity by collecting time-resolved measurements or images with decreased background autofluorescence while eliminating the need for expensive optical hardware, superior resistance to photobleaching, amenability to quantitation, and facile bioconjugation schemes. We isolated rare-earth doped strontium aluminate PLNPs from larger-particle commercial materials by wet milling and differential sedimentation and water-stabilized the particles by silica encapsulation using a modified Stöber process. Surface treatment with aldehyde silane followed by reductive amination with heterobifunctional amine-poly(ethylene glycol)-carboxyl allowed covalent attachment of proteins to the particles using standard carbodiimide chemistry. NeutrAvidin PLNPs were used in lateral flow assays (LFAs) with biotinylated lysozyme as a model analyte in buffer and monoclonal anti-lysozyme HyHEL-5 antibodies at the test line. Preliminary experiments revealed a limit of detection below 100 pg/mL using the NeutrAvidin PLNPs, which was approximately an order of magnitude more sensitive than colloidal gold.

Point-of-care detection of pathogens is medically valuable but poses challenging trade-offs between instrument complexity and clinical and analytical sensitivity. Here we introduce a diagnostic platform utilizing lithographically fabricated micron-scale forms of cubic retroreflectors, arguably one of the most optically detectable human artifacts, as reporter labels for use in sensitive immunoassays. We demonstrate the applicability of this novel optical label in a simple assay format in which retroreflector cubes are first mixed with the sample. The cubes are then allowed to settle onto an immuno-capture surface, followed by inversion for gravity-driven removal of nonspecifically bound cubes. Cubes bridged to the capture surface by the analyte are detected using inexpensive, low-numerical aperture optics. For model bacterial and viral pathogens, sensitivity in 10% human serum was found to be 10(4) bacterial cells/mL and 10(4) virus particles/mL, consistent with clinical utility.

In this work, we demonstrate that signal-masking reagents together with appropriate capture antibody carriers can eliminate the washing steps in sandwich immunoassays. A flotation immunoassay (FI) platform was developed with horseradish peroxidase chemiluminescence as the reporter system, the dye Brilliant Blue FCF as the signal-masking reagent and buoyant silica micro-bubbles as the capture antibody carriers. Only reporters captured on micro-bubbles float above the dye and become visible in an analyte-dependent manner. These FIs are capable of detecting proteins down to attomole levels and as few as 106 virus particles. This signal-masking strategy represents a novel approach to simple, sensitive and quantitative immunoassays in both laboratory and point-of-care settings.

Two types of viral nanoparticles were functionalized with target-specific antibodies and multiple copies of an enzymatic reporter (horseradish peroxidase). The particles were successfully integrated into an immunochromatographic assay detecting MS2 bacteriophage, a model for viral pathogens. The sensitivity of the assay was greatly superior to conventional gold nanoparticle lateral flow assays, and results could easily be evaluated, even without advanced lab instruments.