Universal Detection Scheme Research Articles

An Enzyme- and Label-Free Fluorescence Aptasensor for Detection of Thrombin Based on Graphene Oxide and G-Quadruplex.

An enzyme- and label-free aptamer-based assay is described for the determination of thrombin. A DNA strand (S) consisting of two parts was designed, where the first (Sa) is the thrombin-binding aptamer and the second (Se) is a G-quadruplex. In the absence of thrombin, Sa is readily adsorbed by graphene oxide (GO), which has a preference for ss-DNA rather than for ds-DNA. Upon the addition of the N-methyl-mesoporphyrin IX (NMM), its fluorescence (with excitation/emission at 399/610 nm) is quenched by GO. In contrast, in the presence of thrombin, the aptamer will bind thrombin, and thus, be separated from GO. As a result, fluorescence will be enhanced. The increase is linear in the 0.37 nM to 50 µM thrombin concentration range, and the detection limit is 0.37 nM. The method is highly selective over other proteins, cost-effective, and simple. In our perception, it represents a universal detection scheme that may be applied to other targets according to the proper choice of the aptamer sequence and formation of a suitable aptamer-target pair.

Development and Characterization of a Laser-Induced Acoustic Desorption Source.

A laser-induced acoustic desorption source, developed for use at central facilities, such as free-electron lasers, is presented. It features prolonged measurement times and a fixed interaction point. A novel sample deposition method using aerosol spraying provides a uniform sample coverage and hence stable signal intensity. Utilizing strong-field ionization as a universal detection scheme, the produced molecular plume is characterized in terms of number density, spatial extend, fragmentation, temporal distribution, translational velocity, and translational temperature. The effect of desorption laser intensity on these plume properties is evaluated. While translational velocity is invariant for different desorption laser intensities, pointing to a nonthermal desorption mechanism, the translational temperature increases significantly and higher fragmentation is observed with increased desorption laser fluence.

Chemiluminescence resonance energy transfer-based detection for microchip electrophoresis.

Since the channels in micro- and nanofluidic devices are extremely small, a sensitive detection is required following microchip electrophoresis (MCE). This work describes a highly sensitive and yet universal detection scheme based on chemiluminescence resonance energy transfer (CRET) for MCE. It was found that an efficient CRET occurred between a luminol donor and a CdTe quantum dot (QD) acceptor in the luminol-NaBrO-QD system and that it was sensitively suppressed by the presence of certain organic compounds of biological interest including biogenic amines and thiols, amino acids, organic acids, and steroids. These findings allowed developing sensitive MCE-CL assays for the tested compounds. The proposed MCE-CL methods showed desired analytical figures of merit such as a wide concentration range of linear response. Detection limits obtained were approximately 10(-9) M for biogenic amines including dopamine and epinephrine and approximately 10(-8) M for biogenic thiols (e.g., glutathione and acetylcysteine), organic acids (i.e., ascorbic acid and uric acid), estrogens, and native amino acids. These were 10-1000 times more sensitive than those of previously reported MCE-based methods with chemiluminescence, electrochemical, or laser-induced fluorescence detection for quantifying corresponding compounds. To evaluate the applicability of the present MCE-CL method for analyzing real biological samples, it was used to determine amino acids in individual human red blood cells. Nine amino acids, including Lys, Ser, Ala, Glu, Trp, etc., were detected. The contents ranged from 3 to 31 amol/cell. The assay proved to be simple, quick, reproducible, and very sensitive.

Thermo-optical absorbance detection of native proteins separated by capillary electrophoresis in 10 μm I.D. tubes

Abstract Thermo-optical absorption (TOA) detection of native proteins separated by capillary electrophoresis is demonstrated in 10 μm I.D. tubes. The eluents were on-column pumped at 257 nm and probed by a hologram-based refractive index detector. The use of 10 μm capillaries allowed fast 2-min separations. Slower separations in wider tubes led to limits of detection (LODs) of 7·10−9 M for bovine serum albumin. These LODs are comparable to those obtained with laser induced fluorescence and two orders of magnitude lower than in absorbance detection. Since native fluorescence of proteins is rare, TOA detection appears as a more universal detection scheme and thus suitable for other proteins having smaller amounts of fluorescent amino acids.

Universal chemiluminescence detection using the luminol reaction and the displacement ion effect

A universal detection scheme for ions in solution, based on the stoichiometric displacement of copper(II) ions from a strongly acidic ion-exchange column, is described. The eluted copper(II) ions are detected by their catalytic effect on the chemiluminescent luminol reaction. In conjunction with flow injection, this method of detection is simple, rapid and reproducible, and µg l–1 detection limits can be achieved for mono-, di- and trivalent cations, counter anions and weak acids.