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Investigations on antibody binding to a micro-cantilever coated with a BAM pesticide residue.

Bache M, Taboryski R, Schmid S, Aamand J, Jakobsen MH - Nanoscale Res Lett (2011)

Bottom Line: The stress induced by the binding of a pesticide residue BAM (2,6 dichlorobenzamide) immobilized on a cantilever surface to anti-BAM antibody is measured using the CantiLab4© system from Cantion A/S with four gold-coated cantilevers and piezo resistive readout.The detection mechanism is in principle label-free, but fluorescent-marked antibodies have been used to subsequently verify the binding on the cantilever surface.The system has been analyzed during repeated measurements to investigate whether the CantiLab4© system is a suited platform for a pesticide assay system.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Micro- and Nanotechnology, Technical University of Denmark, DTU Nanotech, Building 345 East, 2800 Kongens Lyngby, Denmark. Michael.bache@nanotech.dtu.dk.

ABSTRACT
The attachment of an antibody to an antigen-coated cantilever has been investigated by repeated experiments, using a cantilever-based detection system by Cantion A/S. The stress induced by the binding of a pesticide residue BAM (2,6 dichlorobenzamide) immobilized on a cantilever surface to anti-BAM antibody is measured using the CantiLab4© system from Cantion A/S with four gold-coated cantilevers and piezo resistive readout. The detection mechanism is in principle label-free, but fluorescent-marked antibodies have been used to subsequently verify the binding on the cantilever surface. The bending and increase in mass of each cantilever has also been investigated using a light interferometer and a Doppler Vibrometer. The system has been analyzed during repeated measurements to investigate whether the CantiLab4© system is a suited platform for a pesticide assay system.

No MeSH data available.


Related in: MedlinePlus

Experimental setup overview. (Above) A schematic overview of the fluidic setup; (Below) Flowchart of the BAM assay on the CantiChip4® system .
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Figure 1: Experimental setup overview. (Above) A schematic overview of the fluidic setup; (Below) Flowchart of the BAM assay on the CantiChip4® system .

Mentions: On an inspected, tested, and clean CantiChip4®, three drops of 0.75 mg/ml BAM-ovalbumine in 1× PBS buffer solution was micro-spotted on cantilever B and C, using a tip voltage of 100 V and pulse width of 20 V. Cantilever A and D was used as reference and was equally micro-spotted with a 1 mg/ml ovalbumine in 1× PBS buffer solution (Figure 1). The chip was incubated overnight in a humidity chamber. A functionalized chip was inserted in the CantiLab4© connected to a fluidic system that consisted of a syringe pump and an 8 channel switchbox. The system was allowed to heat up and stabilize with a continuous flow of 20 μl/min of 1× PBS 0,05% Tween 20 pH 7.4 buffer, for approximately 1 h while a base line was recorded. The experiment consisted of a four-step protocol to minimize false signal sources. The system was first tested against any signal induced by loop switching, second against signal due to buffer injected as a sample. To test for any unspecific antibody attachment signal, a sample of 100 μl of 0.1 mg/ml unspecific mouse Immunoglobulin G (Sigma-Aldrich reagent grade I5381-1 mg, lot.nr.025K7580) Cy5 labeled (Amersham Cy5 Dye™ Antibody monofunctional Labeling Kit) was injected. Following a 5-min buffer flow, finally an injection of 100 μl of the 0.1 mg/ml BAM antibody (Statens Serum Institut, HYB 273-01, Batch nr.03102P01/071008) labeled with Cy3 fluorochrome (Amersham Cy3 Dye™ Antibody mono functional Labeling Kit) was done. Both antibodies were diluted in 1× PBS 0.05%. Tween 20. After the experiment, the Cantion chip was removed from the fluidic chamber and briefly rinsed in Milli-Q water to remove PBS salts. Each test and injection phase of the assay was recorded for 2000 s. The total experiment lasted typically 31/2 h, depending on quality of the output signal and the stabilization period.


Investigations on antibody binding to a micro-cantilever coated with a BAM pesticide residue.

Bache M, Taboryski R, Schmid S, Aamand J, Jakobsen MH - Nanoscale Res Lett (2011)

Experimental setup overview. (Above) A schematic overview of the fluidic setup; (Below) Flowchart of the BAM assay on the CantiChip4® system .
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3211479&req=5

Figure 1: Experimental setup overview. (Above) A schematic overview of the fluidic setup; (Below) Flowchart of the BAM assay on the CantiChip4® system .
Mentions: On an inspected, tested, and clean CantiChip4®, three drops of 0.75 mg/ml BAM-ovalbumine in 1× PBS buffer solution was micro-spotted on cantilever B and C, using a tip voltage of 100 V and pulse width of 20 V. Cantilever A and D was used as reference and was equally micro-spotted with a 1 mg/ml ovalbumine in 1× PBS buffer solution (Figure 1). The chip was incubated overnight in a humidity chamber. A functionalized chip was inserted in the CantiLab4© connected to a fluidic system that consisted of a syringe pump and an 8 channel switchbox. The system was allowed to heat up and stabilize with a continuous flow of 20 μl/min of 1× PBS 0,05% Tween 20 pH 7.4 buffer, for approximately 1 h while a base line was recorded. The experiment consisted of a four-step protocol to minimize false signal sources. The system was first tested against any signal induced by loop switching, second against signal due to buffer injected as a sample. To test for any unspecific antibody attachment signal, a sample of 100 μl of 0.1 mg/ml unspecific mouse Immunoglobulin G (Sigma-Aldrich reagent grade I5381-1 mg, lot.nr.025K7580) Cy5 labeled (Amersham Cy5 Dye™ Antibody monofunctional Labeling Kit) was injected. Following a 5-min buffer flow, finally an injection of 100 μl of the 0.1 mg/ml BAM antibody (Statens Serum Institut, HYB 273-01, Batch nr.03102P01/071008) labeled with Cy3 fluorochrome (Amersham Cy3 Dye™ Antibody mono functional Labeling Kit) was done. Both antibodies were diluted in 1× PBS 0.05%. Tween 20. After the experiment, the Cantion chip was removed from the fluidic chamber and briefly rinsed in Milli-Q water to remove PBS salts. Each test and injection phase of the assay was recorded for 2000 s. The total experiment lasted typically 31/2 h, depending on quality of the output signal and the stabilization period.

Bottom Line: The stress induced by the binding of a pesticide residue BAM (2,6 dichlorobenzamide) immobilized on a cantilever surface to anti-BAM antibody is measured using the CantiLab4© system from Cantion A/S with four gold-coated cantilevers and piezo resistive readout.The detection mechanism is in principle label-free, but fluorescent-marked antibodies have been used to subsequently verify the binding on the cantilever surface.The system has been analyzed during repeated measurements to investigate whether the CantiLab4© system is a suited platform for a pesticide assay system.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Micro- and Nanotechnology, Technical University of Denmark, DTU Nanotech, Building 345 East, 2800 Kongens Lyngby, Denmark. Michael.bache@nanotech.dtu.dk.

ABSTRACT
The attachment of an antibody to an antigen-coated cantilever has been investigated by repeated experiments, using a cantilever-based detection system by Cantion A/S. The stress induced by the binding of a pesticide residue BAM (2,6 dichlorobenzamide) immobilized on a cantilever surface to anti-BAM antibody is measured using the CantiLab4© system from Cantion A/S with four gold-coated cantilevers and piezo resistive readout. The detection mechanism is in principle label-free, but fluorescent-marked antibodies have been used to subsequently verify the binding on the cantilever surface. The bending and increase in mass of each cantilever has also been investigated using a light interferometer and a Doppler Vibrometer. The system has been analyzed during repeated measurements to investigate whether the CantiLab4© system is a suited platform for a pesticide assay system.

No MeSH data available.


Related in: MedlinePlus