Fig 1.
Schematic representation of the magnetic sandwich immunofiltration assay.
A coating antibody (also primary antibody, brown) is immobilized on a porous polyethylene matrix. The Cholera toxin’s subunit B (CTB, green) is captured by the antibody when the CTB-containing solution is flushed over in gravity flow. As the next step, the biotinylated secondary antibody (blue) is flushed over, which also binds to the CTB. Finally the streptavidin-coated superparamagnetic beads (MB, orange) bind to the biotin moiety of the secondary antibody.
Fig 2.
Schematic drawing of an axial cut through the measurement head with schematic view of the working principle.
The measurement head consists of two coils for providing the low driving (f2, yellow, dotted) and high excitation (f1, green, diagonal lines) frequency magnetic field. The differential detection coil is shown with its two compartments for measurement (fine dark blue grid, top) and reference (light blue grid, bottom). The head is designed so that the sample filter (orange) inside the column (gray) is resting at the most sensitive position of the measurement part of the detection coil. Therefore the measurement-coil (dark blue) picks up the MBs signal, as well as, the direct induced signal. The reference-coil (light blue) picks up only the direct induced signal. As they are connected in a differential configuration the resulting signal does not contain these direct induced signals anymore.
Fig 3.
Screenshot of the software to determine the concentration of an unknown sample.
The previously saved fit result for magnetic bead A has been loaded and its calibration points as well as their standard deviation (blue) and the detection limit (red) is displayed. A sample with a spiked concentration of 75 ng/ml was analyzed. A mean value of 6.29 mV with a standard deviation of 0.03 mV was measured and entered. This point is indicated on the calibration curve (black) by green lines and the range is graphically shown with the help of purple lines. For the given values, a concentration of 75.8 ng/ml was found. Together with the standard deviation, this results in a concentration in the range between 75.4 and 76.1 ng/ml.
Fig 4.
Measured magnetic signals of samples with different CTB concentrations.
The mean values and the corresponding standard deviations are depicted as a function of CTB concentration for both types of magnetic beads. The Hill fit curves are also shown.