Time-dependent density functional calculations of optical rotatory dispersion including resonance wavelengths as a potentially useful tool for determining absolute configurations of chiral molecules

The optical rotations for six organic molecules (verbenone, fenchone, camphor, nopinone, Tr?ger's base, dimethyl-cyclopropane) and the transition metal complex [Co(en)(3)](3+) were calculated as a function of wavelength using time-dependent density functional theory (TDDFT). In the calculations, a realistic behavior of the optical rotation in the vicinity of an electronic transition was obtained by using a phenomenological damping parameter of the order of 0.2 eV (0.007 au). In comparison with experiment, for the molecules studied here the sign and order of magnitude of the optical rotation as well as the excitation energies were reasonably well reproduced in most computations. These findings apply to the investigated wavelength ranges typically between about 200 and 650 nm even when using comparatively small basis sets. Such calculations might therefore routinely be applied to help assigning the absolute configurations of chiral molecules. Supplementary calculations of the circular dichroism (CD) and comparison with experimental CD were used for further assessment of the optical rotation calculations. In particular, a combined study of optical rotation and CD turned out to be useful in cases where the optical rotatory dispersion in a specific energy range exhibits a considerable blue or red shift or where it is difficult to reproduce because of an interplay of several competing Cotton effects. The influence of basis set, density functional, and the damping parameter was also investigated.     

Detection of multiple cardiac markers with an integrated acoustic platform for cardiovascular risk assessment

This work describes the application of a newly emerged biosensing configuration incorporating a Surface Acoustic Wave device integrated with a multi-channel microfluidic module for the rapid and efficient analysis of cardiac markers. The examined cardiac markers of creatine kinase MB (CK-MB), cardiac reactive protein (CRP), D-dimer and pregnancy-associated plasma protein A (PAPP-A), comprise a group of both established and emerging heart disease proteins, that has never been probed before with any kind of biochip-related platform. The four markers were successfully detected; kinetics and affinity studies on their interactions with the surface immobilized antibodies are also presented. A concentration detection limit of less than 1 nM was achieved, with a dynamic range of more than two orders of magnitude, covering some of the pathological and healthy areas of interest. Mixtures of biomarkers applied to the device surface were used to prove the specificity of each binding event and investigate the microsystem's performance in the presence of complex fluids, towards future utilization with real samples. The simplicity and multiplexing ability of the integrated platform render the system ideal as a potential diagnostic tool for cardiovascular risk assessment, where simultaneous analysis of various protein markers is required.     

Acoustic determination of polymer molecular weights and rotation times

An acoustic waveguide device was shown to be sensitive to the molecular weight of poly(ethylene glycol) in solution over a molecular weight range determined by the operating frequency of the device. The acoustic device used generates a shear wave with displacement in the plane of the device surface and normal to the direction of propagation. Liquid over the device exhibits viscous coupling to the oscillating surface, affecting propagation of the acoustic wave. The propagation loss was shown to be directly proportional to the weight percentage of the solute. For a given weight percent of polymer in solution, the loss increased with increasing molecular weight until a maximum loss value was reached; this may be due to the fact that rotational times for polymer molecules increase with molecular weight until they reach a point at which the rotation is limited by the oscillation time on the device surface. The molecular weight at which the maximum loss value was attained was 10,000 g/mol for a device operating at 104 MHz and 3350 g/mol for a device operating at 331 MHz, implying a rotational time of 1 ns for each 2200 increase in molecular weight. © 2002 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 40: 1490–1495, 2002     

Convergence of dip-pen nanolithography and acoustic biosensors towards a rapid-analysis multi-sample microsystem

The present work demonstrates for the first time patterning of a ready-to-use biosensor with several different biomolecules using Dip-Pen Nanolithography (DPN) for the development of a procedure towards more rapid and efficient multi-sample detection. The biosensor platform used is based on a Surface Acoustic Wave (SAW) device integrated with a parallel-channel microfluidic module, termed as "microfluidics-on-SAW" ("μF-on-SAW"), for reproducible multi-sample analysis. Lipids with different functionalized head groups were patterned at distinct, microfluidic-formed rectangular domains with sharp edges all located on the same sensor surface; pattern quality was verified using a fluorescent microscope. The functionality of the head groups, the efficiency of the patterning method, and the suitability of DPN for the surface modification of the acoustic device were subsequently examined through acoustic experiments. The μF-on-SAW configuration was used to detect specific binding between the pre-patterned functionalized lipids with their corresponding biomolecules. The achievement of an improved sensitivity (5-fold compared to previous acoustic configurations) and reduced preparation time by at least 2 h clearly indicates the suitability of DPN as a direct patterning method for ready-to-use acoustic sensor devices like the μF-on-SAW towards integrated, rapid-analysis, multi-sample biosensing microsystem development.     

Three-dimensional biomolecule patterning

A new method is demonstrated, where three-dimensional protein structures are made by employing multi-photon polymerization and photobiotin photolysis. The technique enables the construction of arbitrary two- and three-dimensional shapes with submicron resolution. The integrity of the immobilized biotin is confirmed by derivatization with fluorescently labeled streptavidin. Fluorescence microscopy is used in order to visualize the distribution of fluorescent streptavidin on the 3D structure.     

Interaction of cytolytic toxin CytB with a supported lipid bilayer: study using an acoustic wave device

An acoustic technique was used to monitor the interaction of the pore-forming cytolytic toxin CytB with a positively charged supported lipid bilayer. The acoustic device, which is based on a waveguide geometry, is sensitive to changes in the mass of the supported bilayer. The specificity of the interaction, rate and extent of the association, reversibility and effect of previous depositions of toxin were investigated. The CytB was found to bind irreversibly to the lipids at all fractional coverages even when the protein-to-lipid ratio was high enough to imply that the protein was associating with the external surface of the bilayer. The CytB formed stable structures with the bilayer at high protein surface concentrations and did not appear to disrupt the bilayer in the manner of a detergent. The rate of association with the bilayer was found to be directly proportional to the solution concentration of CytB at higher concentrations but appeared to be low at a CytB solution concentration of 5 microg mL(-1), leading to relatively low amounts of CytB being associated with the bilayer.     

Probing mechanical properties of liposomes using acoustic sensors

Acoustic devices were employed to characterize variations in the mechanical properties (density and viscoelasticity) of liposomes composed of 1-oleoyl-2-palmitoyl- sn-glycero-3-phosphocholine (POPC) and cholesterol. Liposome properties were modified in three ways. In some experiments, the POPC/cholesterol ratio was varied prior to deposition on the device surface. Alternatively, the ratio was changed in situ via either insertion of cholesterol or removal of cholesterol with beta-cyclodextrin. This was done for liposomes adsorbed directly on the device surface and for liposomes attached via a biotin-terminated poly(ethylene glycol) linker. The acoustic measurements make use of two simultaneous time-resolved signals: one signal is related to the velocity of the acoustic wave, while the second is related to dissipation of acoustic energy. Together, they provide information not only about the mass (or density) of the probed medium but also about its viscoelastic properties. The cholesterol-induced increase in the surface density of the lipid bilayer was indeed observed in the acoustic data, but the resulting change in signal was larger than expected from the change in surface density. In addition, increasing the bilayer resistance to stretching was found to lead to a greater dissipation of the acoustic energy. The acoustic response is assessed in terms of the possible distortions of the liposomes and the known effects of cholesterol on the mechanical properties of the lipid bilayer that encloses the aqueous core of the liposome. To aid the interpretation of the acoustic response, it is discussed how the above changes in the lipid bilayer will affect the effective viscoelastic properties of the entire liposome/solvent film on the scale of the acoustic wavelength. It was found that the acoustic device is very sensitive to the mechanical properties of lipid vesicles; the response of the acoustic device is explained, and the basic underlying mechanisms of interaction are identified.     

Pulse mode operation of Love wave devices for biosensing applications

In this work we present a novel pulse mode Love wave biosensor that monitors both changes in amplitude and phase. A series of concentrations of 3350 molecular weight poly(ethylene glycol) (PEG) solutions are used as a calibration sequence for the pulse mode system using a network analyzer and high frequency oscilloscope. The operation of the pulse mode system is then compared to the continuous wave network analyzer by showing a sequence of deposition and removal of a model mass layer of palmitoyl-oleoyl-sn-glycerophosphocholine (POPC) vesicles. This experimental apparatus has the potential for making many hundreds of measurements a minute and so allowing the dynamics of fast interactions to be observed.