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To perform the assay, sample DNA is electrophoretically propelled and hybridized to an immobilized DNA probe on the chip and to a fluorophore-labeled DNA probe in solution.
In bridging immunoassays for ADA, interference by the drug target, present in the assay sample, can result in false positive antibody detection.
Thus, the plant system is preferable for the CdS biosynthesis because it does not contain a large amount of the macromolecular compounds that affect the composition of the assay sample.
The assay sample was bound to the tip by pipetting it up and down six times.
Aliquots of the assay sample were then withdrawn with a syringe at specific times and spotted onto nitrocellulose membrane filters.
In both cases, the test material is combined with the assay sample reagent, mixed by hand or vortex, and incubated at room temperature for 15 minutes.
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In addition, saturation of the signal was circumvented by serial dilution of the assay samples.
Using this curve, the amount of ADP in the assay samples was determined.
For the assay, samples were resuspended in 100 µl chloroform; 30 µl were quickly transferred to glass tubes in duplicate and air dried.
Before starting the assay, samples were incubated in 100mmol/L phosphate buffer (pH 7.5) for 1 hour at 37°C to inactivate non-cathepsin S activity, after which the pH was returned to pH 6.0 using 0.5mol/L of MES buffer.
At the end of the assay, samples were counterstained with DAPI for 10 min at RT.
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