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Sensitive and rapid detection of cholera toxin subunit B using magnetic frequency mixing detection
(2019)
Cholera is a life-threatening disease caused by the cholera toxin (CT) as produced by some Vibrio cholerae serogroups. In this research we present a method which directly detects the toxin’s B subunit (CTB) in drinking water. For this purpose we performed a magnetic sandwich immunoassay inside a 3D immunofiltration column. We used two different commercially available antibodies to capture CTB and for binding to superparamagnetic beads. ELISA experiments were performed to select the antibody combination. The beads act as labels for the magnetic frequency mixing detection technique. We show that the limit of detection depends on the type of magnetic beads. A nonlinear Hill curve was fitted to the calibration measurements by means of a custom-written python software. We achieved a sensitive and rapid detection of CTB within a broad concentration range from 0.2 ng/ml to more
than 700 ng/ml.
In modern bioanalytical methods, it is often desired to detect several targets in one sample within one measurement. Immunological methods including those that use superparamagnetic beads are an important group of techniques for these applications. The goal of this work is to investigate the feasibility of simultaneously detecting different superparamagnetic beads acting as markers using the magnetic frequency mixing technique. The frequency of the magnetic excitation field is scanned while the lower driving frequency is kept constant. Due to the particles’ nonlinear magnetization, mixing frequencies are generated. To record their amplitude and phase information, a direct digitization of the pickup-coil’s signal with subsequent Fast Fourier Transformation is performed. By synchronizing both magnetic beads using frequency scanning in magnetic frequency mixing technique magnetic fields, a stable phase information is gained. In this research, it is shown that the amplitude of the dominant mixing component is proportional to the amount of superparamagnetic beads inside a sample. Additionally, it is shown that the phase does not show this behaviour. Excitation frequency scans of different bead types were performed, showing different phases, without correlation to their diverse amplitudes. Two commercially available beads were selected and a determination of their amount in a mixture is performed as a demonstration for multiplex measurements.
The movement of magnetic beads due to a magnetic field gradient is of great interest in different application fields. In this report we present a technique based on a magnetic tweezers setup to measure the velocity factor of magnetically actuated individual superparamagnetic beads in a fluidic environment. Several beads can be tracked simultaneously in order to gain and improve statistics. Furthermore we show our results for different beads with hydrodynamic diameters between 200 and 1000 nm from diverse manufacturers. These measurement data can, for example, be used to determine design parameters for a magnetic separation system, like maximum flow rate and minimum separation time, or to select suitable beads for fixed experimental requirements.
For performing point-of-care molecular diagnostics, magnetic immunoassays constitute a promising alternative to established enzyme-linked immunosorbent assays (ELISA) because they are fast, robust and sensitive. Simultaneous detection of multiple biomolecular targets from one body fluid sample is desired. The aim of this work is to show that multiplex magnetic immunodetection based on magnetic frequency mixing by means of modular immunofiltration columns prepared for different targets is feasible. By calculations of the magnetic response signal, the required spacing between the modules was determined. Immunofiltration columns were manufactured by 3D printing and antibody immobilization was performed in a batch approach. It was shown experimentally that two different target molecules in a sample solution could be individually detected in a single assaying step with magnetic measurements of the corresponding immobilization filters. The arrangement order of the filters and of a negative control did not influence the results. Thus, a simple and reliable approach to multi-target magnetic immunodetection was demonstrated.
With the prevalence of glucosamine- and chondroitin-containing dietary supplements for people with osteoarthritis in the marketplace, it is important to have an accurate and reproducible analytical method for the quantitation of these compounds in finished products. NMR spectroscopic method based both on low- (80 MHz) and high- (500–600 MHz) field NMR instrumentation was established, compared and validated for the determination of chondroitin sulfate and glucosamine in dietary supplements. The proposed method was applied for analysis of 20 different dietary supplements. In the majority of cases, quantification results obtained on the low-field NMR spectrometer are similar to those obtained with high-field 500–600 MHz NMR devices. Validation results in terms of accuracy, precision, reproducibility, limit of detection and recovery demonstrated that the developed method is fit for purpose for the marketed products. The NMR method was extended to the analysis of methylsulfonylmethane, adulterant maltodextrin, acetate and inorganic ions. Low-field NMR can be a quicker and cheaper alternative to more expensive high-field NMR measurements for quality control of the investigated dietary supplements. High-field NMR instrumentation can be more favorable for samples with complex composition due to better resolution, simultaneously giving the possibility of analysis of inorganic species such as potassium and chloride.
Thin films of poly(ethyleneterephthalate) [PET]were exposed to radiation dose ranging from 10 to 30 kGy by using gamma rays in the range 12.8-177.8 MGy using swift light ions of hydrogen. There was no effect of the radiation dose on the optical behaviour of PET as a result of exposure to radiation dose up to 30 kGy brought about by gamma rays but a significant decrease in the optical band gap values was observed when PET was exposed to swift light ions of hydrogen. The data obtained are discussed in terms of optical studies carried out on PET using swift heavy ions.
The purpose of the current study was to examine the reproducibility of fascicle length and pennation angle of gastrocnemius medialis while human walking. To the best of our knowledge, this is the first study of the reproducibility of fascicle length and pennation angle of gastrocnemius medialis in vivo during human gait. Twelve males performed 10 gait trials on a treadmill, in 2 separate days. B-mode ultrasonography, with the ultrasound probe firmly adjusted in the transverse and frontal planes using a special cast, was used to measure the fascicle length and the pennation angle of the gastrocnemius medialis (GM). A Vicon 624 system with three cameras operating at 120 Hz was also used to record the ankle and knee joint angles. The results showed that measurements of fascicle length and pennation angle showed high reproducibility during the gait cycle, both within the same day and between different days. Moreover, the root mean square differences between the repeated waveforms of both variables were very small, compared with their ranges (fascicle length: RMS = ∼3 mm, range: 38–63 mm; pennation angle: RMS = ∼1.5°, range: 22–32°). However, their reproducibility was lower compared to the joint angles. It was found that representative data have to be derived by a wide number of gait trials (fascicle length ∼six trials, pennation angle more than 10 trials), to assure the reliability of the fascicle length and pennation angle in human gait.