Optimised layer systems for immunosensors based on the RIFS transducer

Several approaches to direct affinity sensing are currently under investigation. Prominent approaches based on optical transducers are surface plasmon resonance (SPR), grating couplers and interferometers. In this paper, investigations on the reflectometric interference spectroscopy (RIFS), a transducer based on thin dielectric films, are presented and discussed. The main advantages of the RIFS transducer are its ruggedness, the small active area and simple construction. However limits of detection so far achieved are only in the medium range. One way to improve detection is to increase the signal-to-noise ratio (s/n) in the primary signal (i.e. the reflectance spectra of the transducing film). The influence of the s/n-parameter was investigated by systematic variation of measurement parameters in a given system. A linear relationship between baseline noise of the transducer and s/n ratio of the primary signal was found with an offset value of about 1 pm rms. As the s/n value in the primary signal depends strongly on the optical properties of the transducing layer system, several single and two layer systems have been investigated for theoretical and experimental performance. An up to five-fold increase in signal intensity was achieved as compared to conventional single layer systems. In addition the improved layer systems can be prepared on low refractive index substrates (float glass), producing disposable transducers and giving improved optical systems. The experimental results indicate a thickness resolution of as low as 1 pm rms corresponding to protein coverages below 10 pg/mm². In applications where only moderate detectivity is required, these results offer the possibility for further miniaturisation of the device.     

Nonviral gene delivery using poly‐D/L aspartate‐diethylenetriamine cationic polymers and polyethylene glycol: A two‐step approach

Cationic polymers have received much attention as promising nonviral vectors for gene transfer. However, development of polymers with low cell toxicities and high transfection efficiencies continue to be a significant problem and a major hurdle to their success. Poly‐D /L aspartate‐diethylenetriamine poly(D /L Asp‐DET) polymers were synthesized and evaluated as nonviral gene delivery agents. Poly(D /L Asp‐DET) polymers display endosome buffering capacity. The polymers condense plasmid DNA above N:P ratios of 1 and form polyplex particles of ～50–100 nm, with zeta potentials between neutral and +40 mV. Transmission electron microscopy shows the polyplexes to be uniform in size and shape. Polyplexes maintain the structural integrity of DNA following incubation in nucleases and also show high transfection efficiencies with minimal toxicity in both HCT‐116 and PC‐3 cell culture. However, it is found that these poly(D /L Asp‐DET)/DNA polyplexes immediately aggregate in salt and serum conditions, making them unsuitable for use in vivo. Therefore, the polyplexes were further modified by covalent addition of polyethylene glycol (PEG). Introduction of this second step produces PEG‐polyplexes of uniform size (below 100 nm), with neutral zeta potentials that are also stable in both salt and serum conditions. These results suggest poly(D /L Asp‐DET) cationic polymers as potentially safe and efficient nonviral gene delivery agents. © 2011 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2012     

Characterisation and optimisation of an immunoprobe for triazines

The characterisation and optimisation of an optical immunoassay with label free detection based on Reflectometric Interference Spectroscopy (RIfS) is presented. The immunoprobe is operated in a sequential scheme, where Fab-fragments react with analyte molecules in a first step. In a second step the optical transducer is used to quantify the amount of unoccupied Fab- fragments in the reaction mixture binding to the hapten-modified transducer surface. For optimisation of the test, the Fab-fragment concentration was varied between 2×10^-8 mol/l and 2.5× 10^–9 mol/l. Down to a concentration of 5×10^-9 mol/l a reduction in the limit of detection has been observed. At the lowest concentration investigated no further improvement has been found due to a reduced binding of the analyte and a strong decrease of antibody binding at the transducer surface. This finding could be explained by the thermodynamics of the antigen-antibody reaction and the performance of the optical transducer used. The limit of detection obtained is discussed with respect to thermodynamics, transducer characteristics and immunoprobe test format.     

Variational optimization of the 2DM: approaching three-index accuracy using extended cluster constraints

The reduced density matrix is variationally optimized for the two-dimensional Hubbard model. Exploiting all symmetries present in the system, we have been able to study 6 × 6 lattices at various fillings and different values for the on-site repulsion, using the highly accurate but computationally expensive three-index conditions. To reduce the computational cost we study the performance of imposing the three-index constraints on local clusters of 2 × 2 and 3 × 3 sites. We subsequently derive new constraints which extend these cluster constraints to incorporate the open-system nature of a cluster on a larger lattice. The feasibility of implementing these new constraints is demonstrated by performing a proof-of-principle calculation on the 6 × 6 lattice. It is shown that a large portion of the three-index result can be recovered using these extended cluster constraints, at a fraction of the computational cost.     

Influence of the ferroelastic twin domain structure on the {100} surface morphology of LaAlO3 HTSC substrates

LaAlO₃ crystals have been investigated with differential scanning calorimetry (DSC), high-precision X-ray powder diffraction (XRD) and scanning force microscopy (SFM). The DSC measurements show the second-order phase transition of LaAlO₃ at 544°C, where LaAlO₃ changes its symmetry from the cubic Pm3m high-temperature phase to the pseudocubic rhombohedral low-temperature phase. This paraelastic to improper ferroelastic phase transition causes twinning in the {100} and {110} planes of the pseudocubic lattice. The twin angles between the surface {100}_pseudocubic planes of twin domains were measured by SFM on the surface of a macroscopic (100)_cubic cut crystal plate. The misorientation angle ω₁₀₀ between {100} twins is 0.195(8)°, while {110} twinning gives an angle of ω₁₁₀=0.276(7)°. The two twin kink angles correspond to a rhombohedral angle of the pseudocubic cell of the phase as ₁=90.0973(40)° and ₂=90.0975(30)°, respectively. The XRD result for this rhombohedral angle is =90.096(1)°. The orientation of the misfit steps formed during annealing after mechanical surface polishing depends on the domain orientation and pattern during polishing. Any heating close to or above T_c changes the domain pattern. Footprints of previous domain patterns can thus be found on the surface in the form of surface corrugation and changes in the shape and orientation of misfit steps.     

Label free binding assay with spectroscopic detection for pharmaceutical screening

Many therapeutic drugs exert their effects by interaction with well defined molecular targets. Increasing knowledge in molecular biology allows identification of more and more molecular key compounds and in consequence a molecular approach to disease and therapy. As binding of drugs to their target compounds is a key event, binding assays with an appropriate target molecule are useful means for primary screening of novel substances. We have investigated the potential of thin film interference spectroscopy (RIFS) as a label free detection method for pharmaceutical screening in a binding inhibition assay. To meet the throughput requirements in pharmaceutical screening a parallel detection system based on imaging spectroscopy was constructed. Thrombin/thrombin inhibitor interaction was investigated as a model system. The thin film transducer was covalently modified with a thrombin inhibitor. Specific binding of thrombin and binding inhibition by inhibitor compounds could be observed. A test cycle of less than 10 min could be reached. The parallel setup allows the simultaneous detection of 96 binding curves and can reach a throughput of more than 10⁶ samples per year. Received: 26 November 1996 / Revised: 17 February 1997 / Accepted: 22 February 1997     

Optoacoustic monitoring of blood hemoglobin concentration: a pilot clinical study

The optoacoustic technique is noninvasive, has high spatial resolution, and potentially can be used to measure the total hemoglobin concentration ([THb]) continuously and accurately. We performed in vitro measurements in blood and in vivo tests in healthy volunteers. Our clinical protocol included rapid infusion of intravenous saline to simulate rapid change in the [THb] during fluid therapy or surgery. Optoacoustic measurements were made from the wrist area overlying the radial artery for more than 1 h. The amplitude of the optoacoustic signal generated in the radial artery closely followed the [THb] measured directly in concurrently collected blood samples.