ABSTRACT
Tuberculosis (TB) is the leading global cause of death from a single infectious agent. Registered incidence rates are low, especially in low-resource countries with weak health systems, due to the disadvantages of current diagnostic techniques. A major effort is directed to develop a point-of-care (POC) platform to reduce TB deaths with a prompt and reliable low-cost technique. In the frame of the European POCKET Project, a novel POC platform for the direct and noninvasive detection of TB in human urine was developed. The photonic sensor chip is integrated in a disposable cartridge and is based on a highly sensitive Mach-Zehnder Interferometer (MZI) transducer combined with an on-chip spectral filter. The required elements for the readout are integrated in an instrument prototype, which allows real-time monitoring and data processing. In this work, the novel POC platform has been employed for the direct detection of lipoarabinomannan (LAM), a lipopolysaccharide found in the mycobacterium cell wall. After the optimization of several parameters, a limit of detection of 475 pg/mL (27.14 pM) was achieved using a direct immunoassay in undiluted human urine in less than 15 min. A final validation of the technique was performed using 20 clinical samples from TB patients and healthy donors, allowing the detection of TB in people regardless of HIV coinfection. The results show excellent correlation to those obtained with standard techniques. These promising results demonstrate the high sensitivity, specificity and applicability of our novel POC platform, which could be used during routine check-ups in developing countries.
Subject(s)
Immunoassay/methods , Lipopolysaccharides/urine , Tuberculosis/diagnosis , Antibodies, Immobilized/chemistry , Antibodies, Immobilized/immunology , Humans , Limit of Detection , Lipopolysaccharides/immunology , Mycobacterium tuberculosis/metabolism , Point-of-Care SystemsABSTRACT
Plasmonic nano-apertures are commonly used for the detection of small particles such as nanoparticles and proteins by exploiting electrical and optical techniques. Plasmonic nanopores are metallic nano-apertures sitting on a thin membrane with a tiny hole. It has been shown that plasmonic nanopores with a given geometry identify internal molecules using Surface Enhanced Raman Spectroscopy (SERS). However, label-free identification of a single dielectric nanoparticle requires a highly localized field comparable to the size of the particle. Additionally, the particle's Brownian motion can jeopardize the amount of photons collected from a single particle. Here, we demonstrate that the combination of optical trapping and SERS can be used for the detection and identification of 20 nm polystyrene nanoparticles in plasmonic nanopores. This work is anticipated to contribute to the detection of small bioparticles, optical trapping and nanotribology studies.
ABSTRACT
In this work we present the use of a silicon-on-insulator (SOI) chip featuring an array of 64 optical ring resonators used as refractive index sensors for real-time and label-free DNA detection. Single ring functionalisation was achieved using a click reaction after precise nanolitre spotting of specific hexynyl-terminated DNA capture probes to link to an azido-silanised chip surface. To demonstrate detectability using the ring resonators and to optimise conditions for solid-phase amplification, hybridisation between short 25-mer single stranded DNA (ssDNA) fragments and a complementary capture probe immobilised on the surface of the ring resonators was carried out and detected through the shift in the resonant wavelength. Using the optimised conditions demonstrated via the solid-phase hybridisation, a 144-bp double stranded DNA (dsDNA) was then detected directly using recombinase and polymerase proteins through on-chip target amplification and solid-phase elongation of immobilised forward primers on specific rings, at a constant temperature of 37°C and in less than 60min, achieving a limit of detection of 7.8·10(-13)M (6·10(5) copies in 50µL). The use of an automatic liquid handler injection instrument connected to an integrated resealable chip interface (RCI) allowed programmable multiple injection protocols. Air plugs between different solutions were introduced to prevent intermixing and a proportional-integral-derivative (PID) temperature controller minimised temperature based drifts.
Subject(s)
Biosensing Techniques/methods , DNA/analysis , Biosensing Techniques/instrumentation , Click Chemistry , Computer Systems , DNA Probes , DNA, Bacterial/analysis , DNA, Bacterial/genetics , DNA-Directed DNA Polymerase , Enzymes, Immobilized , Equipment Design , Francisella tularensis/genetics , Immobilized Nucleic Acids , Nanotechnology , Nucleic Acid Amplification Techniques , Nucleic Acid Hybridization , Recombinases , Silicon , Solid-Phase Synthesis TechniquesABSTRACT
A classical 3-port optical circulator is demonstrated on the silicon-on-insulator (SOI) platform. A garnet die with a magneto-optical cerium-doped yttrium iron garnet (Ce:YIG) layer is bonded on top of a Mach-Zehnder interferometer circuit using a thin adhesive bonding layer. The power transmission between different ports is characterized in the presence of an external magnetic field, transversal to the light propagation direction. An isolation of 22 dB is measured at a wavelength of 1562 nm.
