Naked-eye nanobiosensor for therapeutic drug monitoring of methotrexate.

Sensing of methotrexate at clinically-relevant concentrations was achieved with a plasmon-coupling assay. In this assay, free methotrexate and folic acid Au nanoparticles competed for human dihydrofolate reductase (hDHFR)-functionalized Au nanoparticles (Au NP). The hDHFR-functionalized Au NPs were immobilized on a small glass sensor inserted in a portable 4-channel LSPR reader. This allowed rapid (minutes) and sensitive (nanomolar range) measurement of methotrexate concentration by means of total internal reflection plasmonic spectroscopy. The large bathochromic shifts of the plasmon-coupling assay led to striking colour changes visible to the naked eye for methotrexate at clinically-relevant concentrations. The results demonstrate the potential for therapeutic drug monitoring of a widely used chemotherapy agent, as assessed with the naked eye.

Single chip SPR and fluorescent ELISA assay of prostate specific antigen.

A multi-channel system combining fluidics and micropatterned plasmonic materials with wavelength interrogation surface plasmon resonance (SPR) and fluorescence detection was integrated from the combination of a small and motorized fluorescence microscope mounted on a portable 4-channel SPR instrument. The SPR and fluorescent measurements were performed based on the same detection area in a multi-channel fluidic, with a sensing scheme for prostate-specific antigen (PSA) consisting of a sandwich assay with a capture anti-PSA immobilized onto the SPR sensor and a detection anti-PSA modified with horseradish peroxidase (HRP). In this dual-detection instrument, fluorescence was measured from the solution side of the micropatterned gold film, while the interface between the glass prism and the gold film served to interrogate the SPR response. The SPR sensors were comprised of microhole arrays fabricated by photolithography to enhance the instrumental response for PSA detection by approximately a factor of 2 to 3 and they were coated with a self-assembled monolayer of a peptide (3-MPA-HHHDD-OH) to minimize nonspecific adsorption. PSA was successfully detected at clinical concentrations from 10 pM to 50 nM with this integrated system in a single assay lasting 12 minutes, almost centering on the desired range for PSA diagnostic tests (>4 ng mL(-1) or >150 pM). The combination of two robust techniques in a single chip and instrument has led to a simple and effective assay that can be carried out on a small and portable instrument providing rapid biodetection of an important cancer biomarker with a dynamic range of nearly 4 orders of magnitude in the clinical range.

Plasmonic sensors for the competitive detection of testosterone.

The ability to detect small molecules in a rapid and sensitive manner is of great importance in the field of clinical chemistry, and the advancement of novel biosensors is key to realising point-of-care analysis for essential targets. Testosterone is an example of such a small molecule, the detection of which is important in both clinical analysis, and in the sporting industry to prevent doping. As such, a portable, rapid and sensitive test for testosterone would be of great use across a variety of analytical fields. Here we report on a novel method of testosterone analysis, based on a competitive inhibition assay utilising functionalized gold nanoparticles. Two sensing platforms are directly compared for the detection of testosterone based on both classical SPR and LSPR. We provide an in-depth discussion on the optimum surface chemistries needed to create a stable detection conjugate before successfully detecting testosterone using our newly developed portable 4-channel SPR instrument. We provide the first detailed study into the comparison of SPR and LSPR for the analysis of a small molecule, and provide a simple and effective method of testosterone detection that could potentially be extended to a variety of different analytes.

Metal-enhanced fluorescence and FRET on nanohole arrays excited at angled incidence.

The influence of experimental parameters on the performance of plasmonic sensors is of great importance in analytical sciences. The plasmon coupling conditions (angle of incidence, metal composition, laser frequency and excitation/emission properties of fluorophores) were thus investigated for surface plasmon-enhanced fluorescence on metallic nanohole arrays. Optimal fluorescence enhancements were achieved when the plasmon resonance, the excitation laser and the fluorophore's excitation wavelengths were matched. The enhancement of the acceptor emission of a rhodamine 6G(Rh6G)-Quasar670™ FRET pair was achieved on the nanohole arrays by tuning the plasmon wavelength with the maximal overlap of the donor's emission and acceptor excitation. Silver nanohole arrays achieved larger fluorescence enhancement than gold nanohole arrays at 532 nm, while gold nanohole arrays led to larger fluorescence enhancement at 635 nm. These results demonstrate the importance of tuning the plasmon coupling conditions for surface plasmon-enhanced fluorescence sensing.

Low-temperature bitumen stiffness and viscous paraffinic nano- and micro-domains by cryogenic AFM and PDM.

In an effort to better understand the structure and behaviour of bitumen in low temperature, we describe the first use of cryogenic atomic force microscopy and phase detection microscopy to characterize bitumen nano- and micro-structures. The results were interpreted in light of glass transition temperatures (T(g)s) for bitumen fractions. The domains visible by microscopy, the catana, peri and para phases, were attributed to domains rich in asphaltenes, naphthene and polar aromatics, and saturates, respectively. Between -10 degrees C and -30 degrees C, atomic force microscopy images revealed topographic features not visible in atomic force microscopy images acquired at room temperature. According to phase detection microscopy and T(g)s, the features were assigned to viscous unfrozen saturates. Upon cooling to -72 degrees C, unfrozen domains of 20-400 nm were observed. These domains were found in the paraphase rich in saturates and in the periphase rich in naphthene aromatics and polar aromatics. The findings indicate that new viscous domains form upon cooling to low temperatures owing to phase segregation, and that some bitumens are never entirely rigid in low temperatures.

Bitumen morphologies by phase-detection atomic force microscopy.

Summary Bitumen is a complex mixture of hydrocarbons for which microstructural knowledge is incomplete. In an effort to detail this microstructure, 13 bitumens were analysed by phase-detection atomic force microscopy. Based on morphology, the bitumens could be classified into three distinct groups. One group showed fine domains down to 0.1 microm, another showed domains of about 1 microm, and a third group showed up to four different domains or phases of different sizes and shapes. No correlation was found between the atomic force microscopy morphology and the composition based on asphaltenes, polar aromatics, naphthene aromatics and saturates. A high correlation was found between the area of the 'bee-like' structures and the vanadium and nickel content in bitumen, and between the atomic force microscopy groups and the average size of molecular planes made of fused aromatics. The morphology and the molecular arrangements in bitumen thus appear to be partly governed by the molecular planes and the polarity defined by metallic cations.

Sensitive and real-time fiber-optic-based surface plasmon resonance sensors for myoglobin and cardiac troponin I.

A sensor to detect markers of cardiac muscle cell death at less than 3ngml(-1) and in less than 10min has been achieved. This fiber-optic-based surface plasmon resonance (SPR) sensor is being applied to detect myoglobin (MG) and cardiac troponin I (cTnI) in HEPES buffered saline solution. An in vivo sensor for the early detection of the onset of myocardial infarction (MI) will greatly enhance the patient care. MG and cTnI are two biological markers released from dying cardiac muscle cells during an MI, and their detection at biologically-relevant levels can be diagnostic of MI. Antibodies specific to an antigen of interest are attached to a carboxymethylated dextran layer on a gold SPR surface. With the method developed, the lower limit of detection (LOD) for MG is 2.9ngml(-1) at 25 degrees C. The biological level for MG reaches 15-30ngml(-1) in patient blood after myocardial damage. A Langmuir adsorption isotherm describes the binding well. For cTnI, a lower detection limit of 1.4ngml(-1) was achieved in preliminary tests. cTnI levels are in the range of 1-3ngml(-1) in patient blood after myocardial damage. The antibody reaction with the carboxymethylated dextran surface was optimized by modifying the reaction pH, the temperature, and the dextran chain length.

[Upward displacement of the ST segment of an exercise electrocardiogram. Correlation with coronarography]. / Sus-décalage du segment ST à l'électrocardiogramme d'effort Corrélation avec la coronarographie

7 cases with upward displacement of the R-ST segment in the course of effort were compared with the data derived from the coronary arteriography. The lesions were found to be severe in 4 cases, mildly significant or not at all so, in 3 cases. The sub-epicardial lesion on effort, as Prinzmetal's angina, does not make it possible to foresee the degree of the responsible stenosis, which may vary through a superadded spasm, and encourage to undertake an aorta-to-coronary artery by-pass.