A novel ultra‐low‐cost biochemical analysis platform to quantify protein dissociation binding constants and kinetics using paper microfluidics is reported. This approach marries video imaging with ...one of humankind's oldest materials: paper, requiring no large, expensive laboratory equipment, complex microfluidics or external power. Temporal measurements of nanoparticle–antibody conjugates binding on paper is found to follow the Langmuir Adsorption Model. This is exploited to measure a series of antibody–antigen dissociation constants on paper, showing excellent agreement with a gold‐standard benchtop interferometer. The concept is demonstrated with a camera and low‐end smartphone, 500‐fold cheaper than the reference method, and can be multiplexed to measure ten reactions in parallel. These findings will help to widen access to quantitative analytical biochemistry, for diverse applications spanning disease diagnostics, drug discovery, and environmental analysis in resource‐limited settings.
An ultra‐low‐cost platform sensor technology is reported that marries consumer video imaging with nitrocellulose paper to engineer a portable, multiplexed assay to quantify ligand–receptor kinetics. Excellent agreement is found between binding constants measured using video paper analytical devices, and benchtop interferometry.
The combination of two-dimensional materials and metal nanoparticles (MNPs) allows the fabrication of novel nanocomposites with unique physical/chemical properties exploitable in high-performance ...smart devices and biosensing strategies. Current methods to obtain graphene-based films decorated with noble MNPs are cumbersome, poorly reproducible, and difficult to scale up. Herein, we propose a straightforward, versatile, surfactant-free, and single-step technique to produce reduced graphene oxide (rGO) conductive films integrating “naked” noble MNPs. This method relies on the instantaneous laser-induced co-reduction of graphene oxide and metal cations, resulting in highly exfoliated rGO nanosheets embedding gold, silver, and platinum NPs. The production procedure has been optimized, and the obtained nanomaterials are fully characterized; the hybrid nanosheets have been easily transferred onto lab-made screen-printed electrodes preserving their nanoarchitecture. The Au@rGO-, Ag@rGO-, and Pt@rGO-based electrodes have been challenged to detect caffeic acid, nitrite, and hydrogen peroxide in model solutions and real samples. The sensors yielded quantitative responses (R 2 ≥ 0.997) with sub-micromolar limits of detections (LODs ≤ 0.6 μM) for all the analytes, allowing accurate quantification in samples (recoveries ≥ 90%; RSD ≤ 14.8%, n = 3). This single-step protocol which requires low cost and minimal equipment will allow the fabrication of free-standing, MNP-embedded rGO films integrable into a variety of scalable smart devices and biosensors.
The ability to measure the levels of diagnostically relevant proteins, such as antibodies, directly at the point of care could significantly impact healthcare. Thus motivated, we explore here the ...E-DNA “scaffold” sensing platform, a rapid, convenient, single-step means to this end. These sensors comprise a rigid nucleic acid “scaffold” attached via a flexible linker to an electrode and modified on its distal end with a redox reporter and a protein binding “recognition element”. The binding of a targeted protein reduces the efficiency with which the redox reporter approaches the electrode, resulting in an easily measured signal change when the sensor is interrogated voltammetrically. Previously we have demonstrated scaffold sensors employing a range of low molecular weight haptens and linear peptides as their recognition elements. Expanding on this here we have characterized sensors employing much larger recognition elements (up to and including full length proteins) in order to (1) define the range of recognition elements suitable for use in the platform; (2) better characterize the platform’s signaling mechanism to aid its design and optimization; and (3) demonstrate the analytical performance of sensors employing full-length proteins as recognition elements. In doing so we have enlarged the range of molecular targets amenable to this rapid and convenient sensing platform.
The effect of the AuNPs size, ranging from 5 nm to 80 nm, on the electrochemical response of screen-printed carbon electrodes (SPCEs) used as electrochemical transducers is investigated for the first ...time. A simple hydrodynamic modelling and calculation at the nanoscale level is applied so as to find the effect of the size of AuNP upon the electrochemical response. The results show that the best electrochemical response for AuNP suspension for the same concentration of total gold is obtained for the 20 nm sized nanoparticles. It is concluded that the Brownian motions avoid a better response for smaller AuNPs that should in fact be related with the best electrochemical signal due to their higher surface area. Finally, the size effect is studied for AuNPs acting as electroactive labels in an immunosensor that employs magnetic beads as platforms of the bioreactions. The best response for the 5 nm AuNPs in this case is due to the fact that in the immunosensing conditions the Brownian motions are minimized because the AuNPs contact with the electrotransducer surface is induced by the immunoreaction and the fast magnetic collection of the nanoparticles used as antibody labels upon application of a magnetic field.
The majority of lateral flow assays (LFAs) use single‐color optical labels to provide a qualitative naked‐eye detection, however this detection method displays two important limitations. First, the ...use of a single‐color label makes the LFA prone to results misinterpretation. Second, it does not allow the precise modulation of the sensitivity and dynamic range of the test. To overcome these limitations, a ratiometric approach is developed. In particular, using anti‐HIgG functionalized red‐fluorescent quantum dots on the conjugate pad (as target dependent labels) and blue‐fluorescent nanoparticles fixed on the test line (as target independent reporters), it is possible to generate a wide color palette (blue, purple, pink, red) on the test line. It is believed that this strategy will facilitate the development of LFAs by easily adjusting their analytical properties to the needs required by the specific application.
Here the use of target‐dependent and target‐independent fluorescence labels to shift the working range and tune the sensitivity of lateral flow assays are combined. Thanks to this ratiometric approach, both an easier quantification of the test's results by naked eye and a quicker optimization of its analytical performance are achieved.
We have developed a paper and gold nanoparticle (AuNP)-based lab-in-a-syringe (LIS) for immunosensing of biomarkers. This simple diagnostic device features simultaneous sampling and vertical-flow ...operation, which means that unlike typical immunosensors, it does not suffer from any delay between sampling and detection. It can handle large-volume, low-concentration samples for analysis in diverse applications (
e.g.
biomedical, environmental, food,
etc.
). Furthermore, its operating range for sample concentration can be tuned by simply changing the volume of the syringed sample, which enables on-demand limits of detection (LOD). The LIS contains two nitrocellulose pads: the
conjugate pad
(which captures the analyte) and the
detection pad
(which signals the presence of the captured analyte) both embedded into reusable plastic cartridges. We demonstrated its efficiency in detecting human IgG (HIgG) (LOD: 1.0 ng mL
−1
) and prostate-specific antigen (PSA) (spiked urine samples; LOD: 1.9 ng mL
−1
). In the field, the LIS can be used for complete on-site analysis or to obtain partially analyzed samples (AuNPs with captured analyte) for subsequent detailed testing in specialized laboratories.
A novel "lab-in-a-syringe" (LIS) using gold nanoparticles with a simple vertical flow immunosensing operation simultaneously occurring while sampling is developed. This approach avoids the delay between sampling and detection and offers a tunable preconcentration capability.
There is a great demand to develop novel techniques that allow useful and complete monitoring of apoptosis, which is a key factor of several diseases and a target for drug development. Here, we ...present the use of a novel dual electrochemical/optical label for the detection and study of apoptosis. We combined the specificity of Annexin-V for phosphatidylserine, a phospholipid expressed in the outer membrane of apoptotic cells, with the optical and electrochemical properties of quantum dots to create a more efficient label. Using this conjugate we addressed three important issues: (i) we made the labeling of apoptotic cells faster (30 min) and easier; (ii) we fully characterized the samples by common cell biological techniques (confocal laser scanning microscopy, scanning electron microscopy and flow cytometry); and (iii) we developed a fast, cheap and quantitative electrochemical detection method for apoptotic cells with results in full agreement with those obtained by flow cytometry.
A novel-electrochemical Annexin-V-QD conjugate, which can be also used as a classical-optical label, is applied for apoptosis detection in THP-1 cell cultures. The electrochemical detection of AnnV-QD labeled cells gives qualitative and quantitative results in a fast and cost-effective way.
The condensation of 1,1'-ferrocenedimethanol and lipoic acid yields a ferrocene derivative (FcD): 1,1'-ferrocenyl bis(methylene lipoic acid ester), a bipodant linker which was used to aggregate gold ...nanoparticles (AuNPs). The FcD/AuNP aggregations displayed a very well-resolved signal related to the ferrocene oxidation to ferrocenium. The aggregates were found to form an efficient transducing system for a novel concept of immunosensing based on the shifting of the ferrocene oxidation potential. αHIgG/FcD/AuNP or chicken αGIgG/FcD/AuNP bioconjugates are used along with carbon SPEs as an immunosensing platform for the detection of HIgG or GIgG as model analytes. As shown by CV and DPV, the antigens addition triggers significant anodic potential shifts (up to 75 mV) of the ferrocene oxidation peak as an immunoresponse for the Ag-Ab recognition. The results from selectivity and specificity experiments using bovine serum albumin, ovalbumin and α-lactalbumin as potentially interfering proteins to HIgG Ag showed that these potential shifts would be solely due to the Ag-Ab recognition events with excellent selectivity and specificity in the presence of up to 1000-fold of the competitive proteins.
We report the development of a tuneable plasmonic nanochain immunoassay with increased sensitivity over traditional monodisperse nanoparticle lateral flow tests. Our approach takes advantage of the ...unique self-assembling properties of polyamidoamine dendrimers with gold nanoparticles in aqueous media to create one-dimensional nanochains, with a distinct red to blue colour change, attributable to a longitudinal plasmon resonance, which can be readily detected by eye and a digital camera. We optimise and characterise nanochain formation and stability using UV-visible spectroscopy, transmission electron microscopy and dynamic light scattering. As a proof-of-principle we focus on the application of nanochains for point-of-care diagnostics for p24, an important biomarker of early HIV infections and successfully detect p24 with a limit of detection of 5 ng ml
in pseudo-serum, 4 fold more sensitive than comparable studies with gold nanoparticles. These findings and underlying concepts highlight the potential of advanced functional organic-inorganic composite nanomaterials to diagnose infections, with broad applicability to non-communicable diseases.
A handheld wireless optical reader for the detection of infectious diseases at the point of care is presented. The device is battery powered and interfaces with paper-based biosensors for the ...quantification of concentration in biological fluids using colorimetric analysis. The results are wirelessly transmitted to a smartphone. The device has been successfully tested with various concentrations of Influenza A nucleoprotein.
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